Compositions and medical devices comprising anti-microbial particles

ABSTRACT

This invention relates to compositions and medical devices comprising anti-microbial active particles, for inhibiting microbial growth. This invention further provides methods of making such compositions and medical devices.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a Continuation Application from This applicationclaims priority from U.S. Provisional Patent Application No. 62/551,806filed Aug. 30, 2017, U.S. Provisional Patent Application No. 62/551,813filed Aug. 30, 2017, and to U.S. Provisional Patent Application No.62/644,604 filed Mar. 19, 2018, which are hereby incorporated byreference by their entirety.

FIELD OF THE INVENTION

This invention relates to compositions and medical devices comprisinganti-microbial active particles, for inhibiting microbial growth. Thisinvention further provides methods of making such compositions andmedical devices.

BACKGROUND OF THE INVENTION

The overwhelming diversity of bacteria in one individual's skin, gastrointestinal tract and oral cavity is well documented, demonstrating acomplex ecosystem anatomically and dynamically in which poly-microbialbiofilms are the norm.

Biofilms formed on tissues outside and inside the organism are the majorcause of infectious diseases. For example in the oral cavity, biofilmformed on dental hard or soft tissue are the major cause of caries andperiodontal disease (Sbordone L., Bortolaia C., Clin Oral Investig 2003;7:181-8). Bacterial biofilm forms on both natural and artificialsurfaces.

Special attention is paid in recent years to artificial surfacescontacting organisms, as these surfaces lack the epithelial shedding, amajor natural mechanism to combat biofilms, thus biofilm accumulation isbecoming a major source of medical problems that may result in lifethreatening complications. Two major factors influence thesusceptibility of a surface to accumulate bacteria: surface roughnessand the surface-free energy which is a property of the material used.Surface roughness has a higher influence on the adhesion of bacteriathan surface-free energy. In this context, artificial restorativematerials typically have a higher surface roughness than naturalsurfaces, and therefore are more prone to bacterial accumulation.Therefore, the development, of new materials that diminishes biofilmformation is a critical topic.

The ultimate goal of the development of materials with antibiofilmproperties is to improve health and reduce disease occurrence. None ofthe existing medical devices can guarantee immediate and comprehensiveelimination of biofilm or prevention of secondary infection.

F or example, in order to sustain the oral defense, dental materialswith the following antibiofilm properties are sought after; (1)inhibition of initial binding of microorganisms (2) preventing biofilmgrowth, (3) affecting microbial metabolism in the biofilm, (4) killingbiofilm bacteria, and (5) detaching biofilm (Busscher H J, Rinastiti M,Siswomihardjo W, van der Mei H C., J Dent Res, 2010; 89:657-65; Marsh PD. J Dent, 2010:38).

Resin-based composites are complex dental materials that consist of ahydrophobic resin matrix and less hydrophobic filler particles, winchimplies that a resin-based composite surface is never a homogeneousinterface but rather one that produces matrix-rich and filler-poorareas, as well as matrix-poor and filler-rich areas (Ionescu A, WutscherE, Brambilla E, Schneider-Feyrer S, Giessibl F J, Hahnel S.; Eur J OralSci 2012; 120:458-65).

Biofilms on composites can cause surface deterioration. Polishing, aswell as differences in the composition of the resin-based composite, mayhave an impact on biofilm formation on the resin-based composite surface(Ono M. et al., Dent Mater J, 2007; 26:613-22). Surface degradation ofresin composites driven by polishing leads to increased roughness,changes in micro hardness, and filler particle exposure upon exposure tobiofilms in vitro. Furthermore, biofilms on composites can cause surfacedeterioration.

There still remains a need for and it would be advantageous to have anextended variety of anti-microbial active composites, pharmaceuticalcompositions and medical devices which are cost-effective, non-toxic andeasy to apply to contaminated surfaces.

SUMMARY OF THE INVENTION

In one embodiment, this invention is directed to a compositioncomprising anti-microbial particles, wherein the particles comprise:

(i) an inorganic or organic core; and

(ii) an anti-microbial active unit chemically bound to the core;

wherein the anti-microbial active unit is connected directly (via abond) or indirectly (via a third linker) to the core;

wherein the anti-microbial active unit comprises an anti-microbialactive group; and

wherein the number of the anti-microbial active groups per eachanti-microbial active unit is between 1-200.

In one embodiment, this invention is directed to a medical devicecomprising anti-microbial particles, wherein the particles comprise:

(i) an inorganic or organic core; and

(ii) anti-microbial active unit chemically bound to the core;

wherein, the anti-microbial active unit is connected directly (via abond) or indirectly (via a third linker) to the core;

wherein, the anti-microbial active unit comprises an anti-microbialactive group; and wherein the number of the anti-microbial active groupsper each anti-microbial active unit is between 1-200.

In another embodiment, the anti-microbially particle is represented bystructure (1):

whereinthe core is an organic polymer or an inorganic material;L₁ is a first linker or a bond;L₂ is a second linker;L₃ is a third linker or a bond;R₁ and R₁′ are each independently alkyl, terpenoid, cycloalkyl, aryl,heterocycle, alkenyl, alkynyl or any combination thereof;R₂ and R₂′ are each independently alkyl, terpenoid, cycloalkyl, aryl,heterocycle, alkenyl, alkynyl or any combination thereof;R₃ and R₃′ are each independently not present, hydrogen, alkyl,terpenoid moiety, cycloalkyl, aryl, heterocycle, a conjugated alkyl,alkenyl, alkynyl or any combination thereof; wherein if R₃ or R₃′ arenot present, the nitrogen is not charged;X₁ and X₂ is each independently a bond, alkylene, alkenylene, oralkynylene;p defines the density of anti-microbial active unit per one sq nm (nm²)of the core surface, wherein said density is of between 0.01-20anti-microbial units per one sq nm (nm²) of the core surface of theparticle;m is each independently an integer between 0 to 200;n₂ is each independently an integer between 0 to 200;wherein m+n₂≥1; andm is an integer between 1 to 200 and the repeating unit is the same ordifferent.

BRIEF DESCRIPTION OF THE DRAWINGS

The subject matter regarded as the invention is particularly pointed outand distinctly claimed in the concluding portion of the specification.The invention, however, both as to organization and method of operation,together with objects, features, and advantages thereof, may best beunderstood by reference to the following detailed description when readwith the accompanying drawings in which:

FIGS. 1A-1C depict anti-microbial active particle scheme. FIG. 1A: anoligomeric/polymeric backbone per one anti-microbial active unit; FIG.1B: a monomeric backbone per one anti-microbial active unit; and FIG.1C: detailed monomeric unit scheme.

FIG. 2 depicts a representative scheme for the preparation of particlesaccording to this invention wherein the anti-microbial active group is atertiary amine or a quaternary ammonium group comprising at least oneterpenoid moiety and the anti-microbial unit has one monomeric unit (amonomeric backbone, as presented in FIG. 1B); the circles represent theorganic or inorganic core; and R¹—Y—R¹ is a C₁-C₄ alkyl and Y is aleaving group such as halogen or sulfonate.

FIG. 3 depicts a representative scheme for the preparation of a particleof this invention having cinnamyl groups with a core (represented by acircle) via amino-functional linker wherein the anti-microbial unit hasone monomeric unit (a monomeric backbone, as presented in FIG. 1B).Conversion of the tertiary amine to the quaternary ammonium group isoptional, and involves reaction of the tertiary amine with a group R¹—Ywherein R¹ is a C₁-C₄ alkyl and Y is a leaving group such as halogen orsulfonate.

FIG. 4 depicts a representative scheme of three pathways for thepreparation of quaternary ammonium salts (QAS) functionalized particlewherein the anti-microbial unit has one monomeric unit (a monomericbackbone, as presented in FIG. 1B); the circles represents organic orinorganic core. A) by reductive amination to achieve tertiary amine,followed by an alkylation reaction; B) by stepwise alkylation reactions;and C): by reacting a linker functionalized with a leaving group (e.g.,Cl or other halogen) with tertiary amine. R¹ and R² represent C₁-C₄alkyls such as methyl, ethyl, propyl or isopropyl. R¹ and R² may bedifferent or the same group. Y represents any leaving group, for exampleCl, Br or I, or a sulfonate (e.g., mesyl, tosyl).

FIG. 5 depicts schemes of solid support and solution methods for thepreparation of particles of this invention wherein the anti-microbialunit has one monomeric unit (a monomeric backbone, as presented in FIG.1B). The circles represent an organic or inorganic core. Q¹, Q² and Q³are independently selected from the group consisting of ethoxy, methoxy,methyl, ethyl, hydrogen, sulfonate and halide, wherein at least one ofQ¹, Q² and Q³ is a leaving group selected from ethoxy, methoxy,sulfonate (e.g., mesyl, tosyl) and halide. For the sake of clarity thescheme presents a case where Q¹, Q² and Q³ represent leaving groups; Q⁴represents an anti-microbial group; W is selected from the groupconsisting of NH₂, halide, sulfonate and hydroxyl; and n is an integerbetween 1 and 16.

FIG. 6 depicts a representative scheme for the preparation ofdi-cinnamyl groups with core particle (represented as a circle)functionalized utilizing a 12-(triethoxysilyl)-dodecan-1-amine linker byboth solid support method and solution method, wherein theanti-microbial part has one monomeric unit (a monomeric backbone, aspresented in FIG. 1B), n is an integer of 1 to 16.

FIG. 7 depicts a representative scheme for the preparation of particlesaccording to this invention by a solid support method, wherein theanti-microbial unit has an oligomeric or polymeric backbone (more thanone monomeric unit). The circles represent a core. The starting materialis a core terminated on the surface with hydroxyl groups; Q¹⁰¹, Q¹⁰² andQ¹⁰³ and independently alkoxy, alkyl or aryl; LG is Cl, Br, I, mesylate,tosylate or triflate; Hal is Cl, Br or I; q, q¹, q² and q³ areindependently an integer between 0-16; R¹ and R² are each independentlyalkyl, terpenoid, cycloalkyl, aryl, heterocycle, a conjugated alkyl,alkenyl or any combination thereof; and R³ is nothing/not present,hydrogen, alkyl, terpenoid moiety, cycloalkyl, aryl, heterocycle,alkenyl, alkynyl or any combination thereof.

FIGS. 8A-8C depict self-polymerization of trialkoxysilane linker. FIG.8A: self-polymerization of trialkoxysilane linker via solid supportmethod; FIG. 8B: self-polymerization of trialkoxysilane linker insolution; and FIG. 8C: comparison of polymerization of the silane groupsversus simple silanization.

FIG. 9 depicts a representative scheme for the preparation of particlesaccording to this invention in a solution method, wherein theanti-microbial unit has more than one monomeric unit (i.e has anoligomeric or polymeric backbone). The circles represent a core. Thestarting material is a core terminated on the surface with hydroxylgroups; Q¹⁰¹, Q¹⁰² and Q¹⁰³ and independently alkoxy, alkyl or aryl; LGis Cl, Br, I, mesylate, tosylate or triflate; Hal is Cl, Br or I; q, q¹,q² and q³ are independently an integer between 0-16; R¹ and R² are eachindependently alkyl, terpenoid, cycloalkyl, aryl, heterocycle, aconjugated alkyl, alkenyl or any combination thereof; and R³ isnothing/not present, hydrogen, alkyl, terpenoid moiety, cycloalkyl,aryl, heterocycle, alkenyl, alkynyl or any combination thereof.

FIG. 10 depicts a scheme for the preparation of silica basedanti-microbial particles according to this invention comprisingdimethylethylammonium as the anti-microbial active group, in a solidsupport method, wherein the anti-microbial unit has more than onemonomeric unit (i.e has an oligomeric or polymeric backbone).

FIG. 11 depicts a scheme for the preparation of silica basedanti-microbial particles according to this invention comprisingdimethylethylammonium as the anti-microbial active group, in a solutionmethod, wherein the anti-microbial unit has more than one monomeric unit(i.e has an oligomeric or polymeric backbone).

FIG. 12 depicts anti-microbial activity of a polypropylene (PP) matrixwithout and with 1% wt/wt silica based anti-microbial particles (PP+1%NPs) or with 2% wt/wt silica based anti-microbial particles (PP+2% NPs)functionalized with 170 dimethyl octyl ammonium groups per nm²(structure 1; (n₁+n₂)×m×p=170) against the Graham positive bacteriaStaphylococcus aureus (S. aureus). The embedded particles were 186 nm indiameter on average, and the results are compared with the naturalgrowth of S. aureus.

FIG. 13 depicts anti-microbial activity of a polypropylene matrix (PP)without and with 1% wt/wt silica based anti-microbial particles (PP+1%NPs) and with 2% wt/wt silica based anti-microbial particles (PP+2% NPs)functionalized with 170 dimethyl octyl ammonium groups per nm²(structure 1, (n₁+n₂)×m×p=170) against the Graham negative bacteriaPseudomonas aeruginosa (P. aeruginosa). The embedded particles were 186nm in diameter on average, and the results are compared with the naturalgrowth of P. aeruginosa.

FIG. 14 depicts anti-microbial activity of a poly(methyl methacrylate)(PMMA) matrix without and with 1% wt/wt silica based anti-microbialparticles functionalized with 170 dimethyl octyl ammonium groups per nm²(structure 1; (n₁+n₂)×m×p=170) (PMMA+1% NPs), against the Grahamnegative bacteria Pseudomonas aeruginosa (P. aeruginosa). The embeddedparticles were 13 μm in diameter on average, and the results arecompared with the natural growth of P. aeruginosa.

FIG. 15 depicts anti-microbial activity of a poly(methyl methacrylate)matrix (PMMA) without and with 1% wt/wt silica based anti-microbialparticles functionalized with 170 dimethyl octyl ammonium groups per nm²(structure 1; (n₁+n₂)×m×p=170) (PMMA+1% NPs), against the Grahampositive bacteria Staphylococcus aureus (S. aureus). The embeddedparticles were 13 μm in diameter on average, and the results arecompared with the natural growth of S. aureus.

FIG. 16 depicts anti-microbial activity of a poly(methyl methacrylate)matrix without (PMMA) and with silica based anti-microbial particlesfunctionalized with 170 dimethyl octyl ammonium groups per nm²(structure 1; (n₁+n₂)×m×p=170) (PMMA+1% NPs), against the Grahamnegative bacteria Pseudomonas aeruginosa (P. aeruginosa). The embeddedparticles were 186 nm in diameter on average, and the results arecompared with the natural growth of P. aeruginosa.

FIG. 17 depicts anti-microbial activity of a poly(methyl methacrylate)(PMMA) matrix without and with silica based anti-microbial particlesfunctionalized with di-cinnamyl amine groups (PMMA+1% NPs), having amonomeric backbone of:

against the Graham positive bacteria Staphylococcus aureus (S. aureus).The embedded particles were 186 nm in diameter on average and p=4, andthe results are compared with the natural growth of S. aureus.

FIG. 18 depicts the anti-microbial activity of a poly(methylmethacrylate) (PMMA) matrix without and with 1% wt/wt magnetite (Fe₃O₄)based anti-microbial particles (PMMA+1% NPs) or with 2% wt/wt magnetite(Fe₃O₄) based anti-microbial particles (PMMA+2% NPs), functionalizedwith 170 dimethyl octyl ammonium groups per nm² (structure 1,(n₁+n₂)×m×p=170), against the Graham positive bacteria Enterococcusfaecalis (E. faecalis). The embedded particles were 78 nm in diameter onaverage, and the results are compared with the natural growth of E.faecalis.

FIG. 19 depicts the anti-microbial activity of a poly(methylmethacrylate) matrix (PMMA) without and with 2% wt/wt (PMMA+2% NPs) or3% wt/wt (PMMA+3% NPs) silica based anti-microbial particlesfunctionalized with 170 dimethyl octyl ammonium groups per nm²(structure 1; (n₁+n₂)×m×p=170) against the Graham positive bacteriaEnterococcus faecalis (E. faecalis). The embedded particles were 186 nmin diameter on average, and the results are compared with the naturalgrowth of E. faecalis.

FIGS. 20A and 20B: mechanical properties test measuring the young'smodulus of modified polymer including functionalized antibacterialparticles in comparison to unmodified polymer. FIG. 20A: an image of thecylindrical specimens of control (unifast), QSi, and QPEI; FIG. 20B:compressive strength test of modified and unmodified specimens, control:unmodified material (Unifast control), QSi: silica particlesfunctionalized with 170 dimethyl octyl ammonium groups per nm²(structure 1; (n₁+n₂)×m×p=170) and QPEI: quaternary ammoniumpolyethyleneimine.

FIGS. 21A and 21B depicts anti-microbial activity of modified andunmodified specimens of Unifast Trad (a self-cured, methylmethacrylateresin), prepared without (Unifast) or with 8% nanoparticles (NPs):silica+quaternary dimethyl octyl ammonium group (QSi) and PEI+quaternarydimethyl octyl ammonium (QPEI). FIG. 21A: anti-microbial activityagainst the Graham positive bacteria E. faecalis. The results arecompared with the natural growth of E. faecalis. FIG. 21B:anti-microbial activity against the Graham positive bacteria S. aureus.The results are compared with the natural growth of S. aureus.

FIG. 22 presents anti-microbial activity as evaluated by an imprintmethod on blood agar. The samples measured are: 1) dimethylaminefunctionalized silica particles; and 2) tertiary amine with two cinnamylgroups functionalized silica particles.

FIG. 23 depicts the inhibition of E. faecalis bacteria ontopolydimethylsiloxane material, when incorporated 0.5-2% wt/wt of QPEIparticles, (described as PEI in US 2008/0226728 A1).

FIG. 24 depicts anti-microbial activity of a dental composite (example16) with silica based anti-bacterial particles functionalized with 170dimethyl octyl ammonium groups per nm² (structure 1: (n₁+n₂)×m×p=170)against the Graham positive bacteria Enterococcus faecalis (E.faecalis). The embedded particles were 186 nm in diameter on average,and the results are compared with the natural growth of E. faecalis.

FIG. 25 depicts anti-microbial activity of a dental composite (example16) without and with silica based anti-bacterial particlesfunctionalized with di-cinnamyl amine groups (PMMA+1% NPs), having amonomeric backbone,

against the Graham positive bacteria E. faecalis. The embedded particleswere 186 nm in diameter on average and p=4, and the results are comparedwith the natural growth of E. faecalis.

It will be appreciated that for simplicity and clarity of illustration,elements shown in the Figures have not necessarily been drawn to scale.For example, the dimensions of some of the elements may be exaggeratedrelative to other elements for clarity. Further, where consideredappropriate, reference numerals may be repeated among the Figures toindicate corresponding or analogous elements.

DETAILED DESCRIPTION OF THE PRESENT INVENTION

In the following detailed description, numerous specific details are setforth in order to provide a thorough understanding of the invention.However, it will be understood by those skilled in the art that thisinvention may be practiced without these specific details. In otherinstances, well-known methods, procedures, and components have not beendescribed in detail so as not to obscure this invention.

In one embodiment, this invention is directed to a compositioncomprising anti-microbial particles, wherein the particles comprise:

(i) an inorganic or organic core; and

(ii) an anti-microbial active unit chemically bound to the core;

wherein the anti-microbial active unit is connected directly (via abond) or indirectly (via a third linker) to the core;

wherein the anti-microbial active unit comprises an anti-microbialactive group; and

wherein the number of the anti-microbial active groups per eachanti-microbial active unit is between 1-200.

In one embodiment, this invention is directed to a compositioncomprising a polymeric material and anti-microbial particles, whereinthe particles comprise:

(i) an inorganic or organic core; and

(ii) an anti-microbial active unit chemically bound to the core;

wherein the anti-microbial active unit is connected directly (via abond) or indirectly (via a third linker) to the core;

wherein the anti-microbial active unit comprises comprising ananti-microbial active group; and

wherein the number of the anti-microbial active groups per eachanti-microbial active unit is between 1-200.

In one embodiment, this invention is directed to a medical devicecomprising anti-microbial particles, wherein the particles comprise:

(i) an inorganic or organic core; and

(ii) anti-microbial active unit chemically bound to the core;

wherein, the anti-microbial active unit is connected directly (via abond) or indirectly (via a third linker) to the core;

wherein, the anti-microbial active unit comprises an anti-microbialactive group; and wherein the number of the anti-microbial active groupsper each anti-microbial active unit is between 1-200.

Anti-Microbial Particles

In some embodiments, the anti-microbial particles comprise (i) aninorganic or organic core; and (ii) an anti-microbial active partchemically bound to the core. In one embodiment, the anti-microbialactive part comprises one monomeric unit. In one embodiment, theanti-microbial active part comprises more than one monomeric unit. Inanother embodiment, the anti-microbial active part with the more thanone monomeric unit comprises more than one linker. In anotherembodiment, the anti-microbial active unit has between 2-200 monomericunits. In another embodiment, the anti-microbial active unit has between2-5 monomeric units. In another embodiment, the anti-microbial activeunit has between 5-10 monomeric units. In another embodiment, theanti-microbial active unit has between 10-20 monomeric units. In anotherembodiment, the anti-microbial active unit has between 20-50 monomericunits. In another embodiment, the anti-microbial active unit has between50-100 monomeric units. In another embodiment, the anti-microbial activeunit has between 100-200 monomeric units.

In one embodiment, the anti-microbial active unit comprises more thanone monomeric unit. In another embodiment, the monomeric units areconnected to each other via a first linker, a second linker or both. Inanother embodiment, each monomeric unit comprises an anti-microbialactive group. In another embodiment, an anti-microbial active unitcomprises at least one anti-microbial active group. In anotherembodiment, an anti-microbial active unit comprises at least twoanti-microbial active groups. In another embodiment, FIGS. 1A, 1B and 1Cillustrate schematically the anti-microbial active particles of thisinvention (FIG. 1A: more than one monomer; FIG. 1B: one monomeric unitand FIG. 1C: detailed scheme of one monomer).

In some embodiment, the anti-microbial particles are presented bystructure (1):

whereinthe core is an organic polymer or an inorganic material;L₁ is a first linker or a bond;L₂ is a second linker;L₃ is a third linker or a bond;R₁ and R₁′ are each independently alkyl, terpenoid, cycloalkyl, aryl,heterocycle, alkenyl, alkynyl or any combination thereof;R₂ and R₂′ are each independently alkyl, terpenoid, cycloalkyl, aryl,heterocycle, alkenyl, alkynyl or any combination thereof;R₃ and R₃′ are each independently nothing/not present, hydrogen, alkyl,terpenoid moiety, cycloalkyl, aryl, heterocycle, a conjugated alkyl,alkenyl, alkynyl or any combination thereof; wherein if R₃ or R₃′ arenothing/not present, the nitrogen is not charged;X₁ and X₂ is each independently a bond, alkylene, alkenylene, oralkynylene;p defines the density of anti-microbial active units per one sq nm (nm²)of the core surface, wherein said density is of between 0.01-20anti-microbial units per one sq nm (nm²) of the core surface of theparticle;m is each independently an integer between 0 to 200;n₂ is each independently an integer between 0 to 200;wherein m+n₂≥1; andm is an integer between 1 to 200 and the repeating unit is the same ordifferent.

In some embodiments, the anti-microbial particles are represented bystructure (2):

whereinthe core is an organic polymer or an inorganic material;L₁ is a first linker or a bond;L₂ is a second linker;L₃ is a third linker or a bond;R₁ and R₁′ are each independently alkyl, terpenoid, cycloalkyl, aryl,heterocycle, alkenyl, alkynyl or any combination thereof;R₂ and R₂′ are each independently alkyl, terpenoid, cycloalkyl, aryl,heterocycle, alkenyl, alkynyl or any combination thereof;X₁ and X₂ is each independently a bond, alkylene, alkenylene, oralkynylene;p defines the density of anti-microbial active units per one sq nm (nm²)of the core surface, wherein said density is of between 0.01-20anti-microbial units per one sq nm (nm²) of the core surface of theparticle;m is each independently an integer between 0 to 200;n₂ is each independently an integer between 0 to 200;wherein m+n₂≥1;m is an integer between 1 to 200 and the repeating unit is the same ordifferent.

In another embodiment, the number of the anti-microbial active groupsper each anti-microbial active part is at least two, i.e. m+n₂≥2 andm≥1. In another embodiment, the number of the anti-microbial activegroups per each anti-microbial active part is one, i.e. m+n₂=1 and m=1.

In some embodiments, the anti-microbial particles are represented bystructure (3):

whereinthe core is an organic polymer or an inorganic material;L₁ is a first linker or a bond;L₂ is a second linker;L₃ is a third linker or a bond;R₁ and R₁′ are each independently alkyl, terpenoid, cycloalkyl, aryl,heterocycle, alkenyl, alkynyl or any combination thereof;R₂ and R₂′ are each independently alkyl, terpenoid, cycloalkyl, aryl,heterocycle, alkenyl, alkynyl or any combination thereof;X₁ and X₂ is each independently a bond, alkylene, alkenylene, oralkynylene;p defines the density of anti-microbial active units per one sq nm (nm²)of the core surface, wherein said density is of between 0.01-20anti-microbial units per one sq nm (nm²) of the core surface of theparticle;n₁ is each independently an integer between 0 to 200;n₂ is each independently an integer between 0 to 200;wherein n₁+n₂≥1;m is an integer between 1 to 200 and the repeating unit is the same ordifferent.

In another embodiment, the number of the anti-microbial active groupsper each anti-microbial active part is at least two, i.e. n₁+n₂≥2 andm≥1. In another embodiment, the number of the anti-microbial activegroups per each anti-microbial active part is one, i.e. n₁+n₂=1 and m=1.

In another embodiment, the particles of structures (1) to (3) compriseone monomeric unit per one anti-microbial active unit. In anotherembodiment, the particles of structures (1) to (3) comprise more thanone anti-microbial active group per one anti-microbial active unit.

In some embodiments, the anti-microbial particles are represented bystructure (4):

whereinthe core is an organic polymer or an inorganic material;L₁ is a first linker or a bond;L₃ is a third linker or a bond;R₁ is alkyl, terpenoid, cycloalkyl, aryl, heterocycle, alkenyl, alkynylor any combination thereof;R₂ is alkyl, terpenoid, cycloalkyl, aryl, heterocycle, alkenyl, alkynylor any combination thereof;R₃ is nothing/not present, hydrogen, alkyl, terpenoid moiety,cycloalkyl, aryl, heterocycle, alkenyl, alkynyl or any combinationthereof, wherein if R₃ or R₃′ are nothing/not present, the nitrogen isnot charged;X is a bond, alkyl, alkenyl, or alkynyl;X′ is nothing or hydrogen; andp defines the density of anti-microbial active units per one sq nm (nm²)of the core surface, wherein said density is of between 0.01-20anti-microbial units per one sq nm (nm²) of the core surface of theparticle;wherein if L₁ and X are bonds, then the nitrogen is part of the core;wherein at least one of R₁, R₂, R₃ is hydrophobic.

In some embodiments, the anti-microbial particles are represented bystructure (5):

whereinthe core is an organic polymeric material or an inorganic material;L₁ is a first linker or a bond;L₃ is a third linker or a bond;R₁ is alkyl, terpenoid, cycloalkyl, aryl, heterocycle, alkenyl, alkynylor any combination thereof;R₂ is alkyl, terpenoid, cycloalkyl, aryl, heterocycle, alkenyl, alkynylor any combination thereof;X is a bond, alkyl, alkenyl or alkynyl;X′ is nothing or hydrogen; andp defines the density of anti-microbial active units per one sq nm (nm²)of the core surface, wherein said density is of between 0.01-20anti-microbial units per one sq nm (nm²) of the core surface of theparticle;wherein if L₁ and X are bonds, then the nitrogen is an integral part ofthe core;wherein at least one of R₁, R₂ is hydrophobic.

In some embodiments, the anti-microbial particles are represented bystructure (6):

whereinthe core is an organic polymeric material or an inorganic material;L₁ is a first linker or a bond;L₃ is a third linker or a bond;R₁ is alkyl, terpenoid, cycloalkyl, aryl, heterocycle, alkenyl, alkynylor any combination thereof;R₂ is alkyl, terpenoid, cycloalkyl, aryl, heterocycle, alkenyl, alkynylor any combination thereof;X is a bond, alkyl, alkenyl, or alkynyl;X′ is nothing or hydrogen; andp defines the density of anti-microbial active units per one sq nm (nm²)of the core surface, wherein said density is of between 0.01-20anti-microbial units per one sq nm (nm²) of the core surface of theparticle;wherein if L₁ and X are bonds, then the nitrogen is an integral part ofthe core; wherein at least one of R₁, R₂ is hydrophobic.

Specific examples of anti-microbial particles of this invention include:

where n=1-200; “SNP” refers to the a silica core of the particles ofthis invention; and p defines the density of anti-microbial active unitsper one sq nm (nm²) of the core surface, wherein said density is ofbetween 0.01-20 anti-microbial units per one sq nm (nm²) of the coresurface of the particle. In another embodiment, n=1-3. In anotherembodiment, n=3-20. In another embodiment, n=20-50. In anotherembodiment, n=50-100. In another embodiment, n=100-200.

In some embodiments, the anti-microbial particles of structures (1) to(3) a hydrogen may or may not be present at the end of theanti-microbial active unit.

In some embodiments, the term “anti-microbial active group” and the term“monomeric anti-microbial active group” refer to the same and comprise aprotonated tertiary amine, a tertiary amine or a quaternary ammonium, asrepresented by the following formulas:

wherein:R₁ is alkyl, terpenoid, cycloalkyl, aryl, heterocycle, alkenyl, alkynylor any combination thereof;R₂ is alkyl, terpenoid, cycloalkyl, aryl, heterocycle, alkenyl, alkynylor any combination thereof;R₃ is nothing, hydrogen, alkyl, terpenoid moiety, cycloalkyl, aryl,heterocycle, alkenyl, alkynyl or any combination thereof; wherein if R₃or R₃′ are nothing, the nitrogen is not charged.

In another embodiment, at least one of R₁, R₂ or R₃ is hydrophobic.

In another embodiment, the number of the anti-microbial active groupsper each anti-microbial active unit is at least two, i.e. n₁+n₂≥2 andm≥1. In another embodiment, the number of the anti-microbial activegroups per each anti-microbial active unit is one, i.e. n₁+n₂=1 and m=1.

In another embodiment, the particles of structures (4) to (6) compriseone monomeric unit per one anti-microbial active unit. In anotherembodiment, the particles of structures (1) to (3) comprise one or morethan one anti-microbial active group per one anti-microbial active unit.

The anti-microbial active groups of this invention are chemically boundto the core at a surface density of at least one anti-microbial activegroup per 10 sq. nm of the core surface. In another embodiment at least1 anti-microbial group per 1 sq nm of the core surface. In anotherembodiment between 0.001-300 anti-microbial groups per sq nm of the coresurface. In another embodiment between 0.001-250 anti-microbial groupsper sq nm of the core surface. In another embodiment between 0.001-200anti-microbial groups per sq nm of the core surface. In anotherembodiment between 0.001-150 anti-microbial groups per sq nm of the coresurface. In another embodiment between 0.001-100 anti-microbial groupsper sq nm of the core surface. In another embodiment between 0.001-50anti-microbial groups per sq nm of the core surface. In anotherembodiment between 0.001-20 anti-microbial groups per sq nm of the coresurface. In another embodiment between 0.001-17 anti-microbial groupsper sq nm of the core surface. In another embodiment between 0.001-15anti-microbial groups per sq nm of the core surface. In anotherembodiment between 0.001-10 anti-microbial groups per sq nm of the coresurface. In another embodiment between 0.001-4 anti-microbial groups persq nm of the core surface. In another embodiment between 0.001-1anti-microbial groups per sq nm of the core surface. In anotherembodiment between 50-100 anti-microbial groups per sq nm of the coresurface. In another embodiment between 100-150 anti-microbial groups persq nm of the core surface. In another embodiment between 150-200anti-microbial groups per sq nm of the core surface. In anotherembodiment between 200-250 anti-microbial groups per sq nm of the coresurface. In another embodiment between 250-300 anti-microbial groups persq nm of the core surface. In another embodiment between 1-4anti-microbial groups per sq nm of the core surface. In anotherembodiment between 1-6 anti-microbial groups per sq nm of the coresurface. In another embodiment between 1-20 anti-microbial groups per sqnm of the core surface. In another embodiment between 1-10anti-microbial groups per sq nm of the core surface. In anotherembodiment between 1-15 anti-microbial groups per sq nm of the coresurface.

In some embodiments, the number of the anti-microbial active groups[(n₁+n₂)×m] per each anti-microbial active unit is between 1-200. Inanother embodiment, the number of the anti-microbial active groups pereach anti-microbial active unit is between 1-150. In another embodiment,the number of the anti-microbial active groups per each anti-microbialactive unit is between 1-100. In another embodiment, the number of theanti-microbial active groups per each anti-microbial active unit isbetween 1-50. In another embodiment, the number of the anti-microbialactive groups per each anti-microbial active unit is between 1-30. Inanother embodiment, the number of the anti-microbial active groups pereach anti-microbial active unit is between 1-20. In another embodiment,the number of the anti-microbial active groups per each anti-microbialactive unit is between 1-10. In another embodiment, the number of theanti-microbial active groups per each anti-microbial active unit isbetween 50-100. In another embodiment, the number of the anti-microbialactive groups per each anti-microbial active unit is between 100-150. Inanother embodiment, the number of the anti-microbial active unit pereach anti-microbial active unit is between 150-200.

In some embodiments, the number of the monomeric units per eachanti-microbial active unit is between 1-200. In another embodiment, thenumber of the monomeric units per each anti-microbial active unit isbetween 1-150. In another embodiment, the number of the monomeric unitsper each anti-microbial active unit is between 1-100. In anotherembodiment, the number of the monomeric units per each anti-microbialactive unit is between 1-50. In another embodiment, the number of themonomeric units per each anti-microbial active unit is between 1-30. Inanother embodiment, the number of monomeric units per eachanti-microbial active unit is between 1-20. In another embodiment, thenumber of the monomeric units per each anti-microbial active unit isbetween 1-10. In another embodiment, the number of the monomeric unitsper each anti-microbial active unit is between 50-100. In anotherembodiment, the number of the monomeric units per each anti-microbialactive unit is between 100-150. In another embodiment, the number of themonomeric units per each anti-microbial active unit is between 150-200.

In another embodiment, the particle of structures (1) to (6) has aninorganic core. In another embodiment, the particle of structure (1) to(6) has an organic core. In another embodiment, the organic core is apolymeric organic core. In another embodiment, the core is inert. In oneembodiment, the particles of this invention represented by structures(1)-(3) comprise an anti-microbial active group of —⁺N(R₁)(R₂)(R₃),—⁺NH(R₁)(R₂), —N(R₁)(R₂) —⁺N(R₁′)(R₂′)(R₃′), —⁺NH(R₁′)(R₂′) or—N(R₁′)(R₂′). In one embodiment R₁ and/or R₁′, R₂ and/or R₂′ and R₃and/or R₃′ are the same or different and are independently alkyl,terpenoid, cycloalkyl, aryl, heterocycle, alkenyl, alkynyl or anycombination thereof. In another embodiment, R₁, R₂ and R₃ areindependently an alkyl. In another embodiment, R₁ and/or R₁′, R₂ and/orR₂′ and R₃ and/or R₃′ are independently a terpenoid. In anotherembodiment, R₁ and/or R₁′, R₂ and/or R₂′ and R₃ and/or R₃′ areindependently a cycloalkyl. In another embodiment, R₁ and/or R₁′, R₂and/or R₂′ and R₃ and/or R₃′ are independently an aryl. In anotherembodiment, R₁ and/or R₁′, R₂ and/or R₂′ and R₃ and/or R₃′ areindependently a heterocycle. In another embodiment, R₁ and/or R₁′, R₂and/or R₂′ and R₃ and/or R₃′ are independently an alkenyl. In anotherembodiment, R₁ and/or R₁′, R₂ and/or R₂′ and R₃ and/or R₃′ areindependently an alkynyl. In another embodiment, R₃ is nothing. Inanother embodiment, R₃ and/or R₃′ is hydrogen. In another embodiment atleast one of R₁ and/or R₁′, R₂ and/or R₂′ and R₃ and/or R₃′ ishydrophobic alkyl, terpenoid, cycloalkyl, aryl, heterocycle, alkenyl,alkynyl or any combination thereof. Each represents a separateembodiment of this invention.

In another embodiment R₁ and R₁′ are the same. In another embodiment R₂and R₂′ are the same. In another embodiment R₃ and R₃′ are the same. Inanother embodiment R₁ and R₁′ are different. In another embodiment R₂and R₂′ are different. In another embodiment R₃ and R₃′ are different.

As used herein, the term “alkyl” or “alkylene” refer to any linear- orbranched-chain alkyl group containing up to about 24 carbons unlessotherwise specified. In one embodiment, an alkyl includes C₁-C₃ carbons.In one embodiment, an alkyl includes C₁-C₄ carbons. In one embodiment,an alkyl includes C₁-C₅ carbons. In another embodiment, an alkylincludes C₁-C₆ carbons. In another embodiment, an alkyl includes C₁-C₈carbons. In another embodiment, an alkyl includes C₁-C₁₀ carbons. Inanother embodiment, an alkyl includes C₁-C₁₂ carbons. In anotherembodiment, an alkyl includes C₄-C₈ carbons. In another embodiment, analkyl includes C₄-C₁₀ carbons. In another embodiment, an alkyl includeC₄-C₁₈ carbons. In another embodiment, an alkyl include C₄-C₂₄ carbons.In another embodiment, an alkyl includes C₁-C₁₈ carbons. In anotherembodiment, an alkyl includes C₂-C₁₈ carbons. In another embodiment,branched alkyl is an alkyl substituted by alkyl side chains of 1 to 5carbons. In one embodiment, the alkyl group may be unsubstituted. Inanother embodiment, the alkyl group may be substituted by a halogen,haloalkyl, hydroxyl, alkoxy, carbonyl, amido, alkylamido, dialkylamido,cyano, nitro, CO₂H, amino, alkylamino, dialkylamino, carboxyl, thioand/or thioalkyl. In another embodiment hydrophobic alkyl refers to analkyl having at least four carbons. In another embodiment hydrophobicalkyl refers to a C₄-C₂₄ alkyl. In another embodiment hydrophobic alkylrefers to a C₄-C₈ alkyl. In another embodiment hydrophobic alkyl refersto a C₄ alkyl. In another embodiment hydrophobic alkyl refers to a C₅alkyl. In another embodiment hydrophobic alkyl refers to a C₆ alkyl. Inanother embodiment hydrophobic alkyl refers to a C₇ alkyl. In anotherembodiment hydrophobic alkyl refers to a C₈ alkyl.

As used herein, the term “aryl” refers to any aromatic ring that isdirectly bonded to another group and can be either substituted orunsubstituted. The aryl group can be a sole substituent, or the arylgroup can be a component of a larger substituent, such as in anarylalkyl, arylamino, arylamido, etc. Exemplary aryl groups include,without limitation, phenyl, tolyl, xylyl, furanyl, naphthyl, pyridinyl,pyrimidinyl, pyridazinyl, pyrazinyl, triazinyl, thiazolyl, oxazolyl,isooxazolyl, pyrazolyl, imidazolyl, thiophene-yl, pyrrolyl,phenylmethyl, phenylethyl, phenylamino, phenylamido, etc. Substitutionsinclude but are not limited to: F, Cl, Br, I, C₁-C₅ linear or branchedalkyl, C₁-C₅ linear or branched haloalkyl, C₁-C₅ linear or branchedalkoxy, C₁-C₅ linear or branched haloalkoxy, CF₃, CN, NO₂, —CH₂CN, NH₂,NH-alkyl, N(alkyl)₂, hydroxyl, —OC(O)CF₃, —OCH₂Ph, —NHCO-alkyl, COOH,—C(O)Ph, C(O)O-alkyl, C(O)H, or —C(O)NH₂. In another embodimenthydrophobic aryl refers to aryl having at least six carbons.

The term “alkenyl” or “alkenylene” refer to a substance that includes atleast two carbon atoms and at least one double bond. In one embodiment,the alkenyl has 2-7 carbon atoms. In another embodiment, the alkenyl has2-12 carbon atoms. In another embodiment, the alkenyl has 2-10 carbonatoms. In another embodiment, the alkenyl has 3-6 carbon atoms. Inanother embodiment, the alkenyl has 2-4 carbon atoms. In anotherembodiment, the alkenyl has 4-8 carbon atoms. In another embodimenthydrophobic alkenyl refers to alkenyl having at least four carbons. Inanother embodiment hydrophobic alkenyl refers to a C₄-C₈ alkenyl.

The term “alkynyl” or “alkynylene” refers to a substance that includesat least two carbon atoms and at least one triple bond. In oneembodiment, the alkynyl has 2-7 carbon atoms. In another embodiment, thealkynyl has 2-12 carbon atoms. In another embodiment, the alkynyl has2-10 carbon atoms. In another embodiment, the alkynyl has 3-6 carbonatoms. In another embodiment, the alkynyl has 2-4 carbon atoms. Inanother embodiment, the alkynyl has 3-6 carbon atoms. In anotherembodiment, the alkynyl has 4-8 carbon atoms. In another embodimenthydrophobic alkynyl refers to alkynyl having at least four carbons. Inanother embodiment hydrophobic alkynyl refers to a C₄-C₈ alkenyl.

The term “alkoxy” refers in one embodiment to an alky as defined abovebonded to an oxygen. Non limiting examples of alkoxy groups include:methoxy, ethoxy and isopropoxy.

A “cycloalkyl” group refers, in one embodiment, to a ring structurecomprising carbon atoms as ring atoms, which may be either saturated orunsaturated, substituted or unsubstituted. In another embodiment thecycloalkyl is a 3-12 membered ring. In another embodiment the cycloalkylis a 6 membered ring. In another embodiment the cycloalkyl is a 5-7membered ring. In another embodiment the cycloalkyl is a 3-8 memberedring. In another embodiment, the cycloalkyl group may be unsubstitutedor substituted by a halogen, alkyl, haloalkyl, hydroxyl, alkoxy,carbonyl, amido, alkylamido, dialkylamido, cyano, nitro, CO₂H, amino,alkylamino, dialkylamino, carboxyl, thio and/or thioalkyl. In anotherembodiment, the cycloalkyl ring may be fused to another saturated orunsaturated cycloalkyl or heterocyclic 3-8 membered ring. In anotherembodiment, the cycloalkyl ring is a saturated ring. In anotherembodiment, the cycloalkyl ring is an unsaturated ring. Non-limitingexamples of a cycloalkyl group comprise cyclohexyl, cyclohexenyl,cyclopropyl, cyclopropenyl, cyclopentyl, cyclopentenyl, cyclobutyl,cyclobutenyl, cycloctyl, cycloctadienyl (COD), cycloctaene (COE) etc. Inanother embodiment hydrophobic cycloalkyl refers to a cycloalkyl havingat least six carbons.

A “heterocycle” group refers, in one embodiment, to a ring structurecomprising in addition to carbon atoms, sulfur, oxygen, nitrogen or anycombination thereof, as part of the ring. In another embodiment theheterocycle is a 3-12 membered ring. In another embodiment theheterocycle is a 6 membered ring. In another embodiment the heterocycleis a 5-7 membered ring. In another embodiment the heterocycle is a 3-8membered ring. In another embodiment, the heterocycle group may beunsubstituted or substituted by a halogen, alkyl, haloalkyl, hydroxyl,alkoxy, carbonyl, amido, alkylamido, dialkylamido, cyano, nitro, CO₂H,amino, alkylamino, dialkylamino, carboxyl, thio and/or thioalkyl. Inanother embodiment, the heterocycle ring may be fused to anothersaturated or unsaturated cycloalkyl or heterocyclic 3-8 membered ring.In another embodiment, the heterocyclic ring is a saturated ring. Inanother embodiment, the heterocyclic ring is an unsaturated ring. Nonlimiting examples of a heterocyclic rings comprise pyridine, piperidine,morpholine, piperazine, thiophene, pyrrole, benzodioxole, or indole. Inanother embodiment hydrophobic heterocyclic group refers to aheterocycle having at least six carbons.

In one embodiment, at least one of R₁, R₂ and R₃ and/or at least one ofR₁′, R₂′ and R₃′ of structure (1) is hydrophobic. In one embodiment, atleast one of R₁ and R₂ and/or at least one of R₁′ and R₂′ of structures(2) and (3) is hydrophobic.

The term “hydrophobic” refers to an alkyl, alkenyl or alkynyl having atleast four carbons, or the term hydrophobic refers to terpenoid, tocycloalkyl, aryl or heterocycle having at least six carbons. Eachpossibility represents a separate embodiment of this invention

In one embodiment, at least one of R₁, R₂ and R₃ and/or at least one ofR₁′, R₂′ and R₃′ of structure (1) is a C₄-C₂₄ alkyl, C₄-C₂₄ alkenyl,C₄-C₂₄ alkynyl or a terpenoid. In one embodiment, at least one of R₁ andR₂ and/or at least one of R₁′ and R₂′ of structures (2) and (3) is aC₄-C₂₄ alkyl, C₄-C₂₄ alkenyl, C₄-C₂₄ alkynyl or a terpenoid. Eachpossibility represents a separate embodiment of this invention.

In one embodiment, at least one of R₁, R₂ and R₃ and/or at least one ofR₁′, R₂′ and R₃′ of structure (1) is a C₄-C₈ alkyl, C₄-C₈ alkenyl, C₄-C₈alkynyl or a terpenoid. In one embodiment, at least one of R₁ and R₂and/or at least one of R₁′ and R₂′ of structures (2) and (3) is a C₄-C₈alkyl, C₄-C₈ alkenyl, C₄-C₈ alkynyl or a terpenoid. Each possibilityrepresents a separate embodiment of this invention.

In one embodiment, R₁ and/or R₁′ of structures (1) to (6) is aterpenoid. In another embodiment, R₁ and/or R₁′ is a terpenoid and R₂and/or R₂′ is a C₁-C₄ alkyl. In another embodiment, the core is anorganic polymeric core, R₃ and/or R₃′ is nothing and R₁ and/or R₁′ is aterpenoid. In another embodiment, the core is an organic polymeric core,R₃ and/or R₃′ is a hydrogen and R₁ and/or R₁′ is a terpenoid. In anotherembodiment, the core is an inorganic core, R₃ and/or R₃′ is nothing andR₁ and/or R₁′ is a terpenoid. In another embodiment, the core is aninorganic core, R₃ and/or R₃′ is a hydrogen and R₁ and/or R₁′ is aterpenoid. In another embodiment, the core is an inorganic core, R₃and/or R₃′ is a C₁-C₂₄ alkyl, terpenoid, cycloalkyl, aryl, heterocycle,a conjugated C₁-C₂₄ alkyl, C₁-C₂₄ alkenyl, C₁-C₂₄ alkynyl or anycombination thereof and R₁ and/or R₁′ is a terpenoid.

In one embodiment “p” of structures (1) to (6) defines the surfacedensity of the anti-microbial active units per 1 sq nm of the coresurface. In another embodiment “p” is, between 0.01-30 anti-microbialactive units per 1 sq nm of the core surface. In another embodiment “p”is, between 0.01-20 anti-microbial active units per 1 sq nm of the coresurface. In another embodiment “p” is, between 0.01-10 anti-microbialactive units per 1 sq nm of the core surface. In another embodiment “p”is, between 0.01-15 anti-microbial active units per 1 sq nm of the coresurface. In another embodiment “p” is, between 0.01-5 anti-microbialactive units per 1 sq nm of the core surface.

In one embodiment, n₁ of structures (1) to (6) is between 0-200. Inanother embodiment, n₁ is between 0-10. In another embodiment, n₁ isbetween 10-20. In another embodiment, n₁ is between 20-30. In anotherembodiment, n₁ is between 30-40. In another embodiment, n₁ is between40-50. In another embodiment, n₁ is between 50-60. In anotherembodiment, n₁ is between 60-70. In another embodiment, n₁ is between70-80. In another embodiment, n₁ is between 80-90. In anotherembodiment, n₁ is between 90-100. In another embodiment, n₁ is between100-110. In another embodiment, n₁ is between 110-120. In anotherembodiment, n₁ is between 120-130. In another embodiment, n₁ is between130-140. In another embodiment, n₁ is between 140-150. In anotherembodiment, n₁ is between 150-160. In another embodiment, n₁ is between160-170. In another embodiment, n₁ is between 170-180. In anotherembodiment, n₁ is between 180-190. In another embodiment, n₁ is between190-200.

In one embodiment, n₂ of structures (1) to (6) is between 0-200. Inanother embodiment, n₂ is between 0-10. In another embodiment, n₂ isbetween 10-20. In another embodiment, n₂ is between 20-30. In anotherembodiment, n₂ is between 30-40. In another embodiment, n₂ is between40-50. In another embodiment, n₂ is between 50-60. In anotherembodiment, n₂ is between 60-70. In another embodiment, n₂ is between70-80. In another embodiment, n₂ is between 80-90. In anotherembodiment, n₂ is between 90-100. In another embodiment, n₂ is between100-110. In another embodiment, n₂ is between 110-120. In anotherembodiment, n₂ is between 120-130. In another embodiment, n₂ is between130-140. In another embodiment, n₂ is between 140-150. In anotherembodiment, n₂ is between 150-160. In another embodiment, n₂ is between160-170. In another embodiment, n₂ is between 170-180. In anotherembodiment, n₂ is between 180-190. In another embodiment, n₂ is between190-200.

In one embodiment, m of structures (1) to (6) is between 1-200. Inanother embodiment, m is between 1-10. In another embodiment, m isbetween 10-20. In another embodiment, m is between 20-30. In anotherembodiment, m is between 30-40. In another embodiment, m is between40-50. In another embodiment, m is between 50-60. In another embodiment,m is between 60-70. In another embodiment, m is between 70-80. Inanother embodiment, m is between 80-90. In another embodiment, m isbetween 90-100. In another embodiment, m is between 100-110. In anotherembodiment, m is between 110-120. In another embodiment, m is between120-130. In another embodiment, m is between 130-140. In anotherembodiment, m is between 140-150. In another embodiment, m is between150-160. In another embodiment, m is between 160-170. In anotherembodiment, m is between 170-180. In another embodiment, m is between180-190. In another embodiment, m is between 190-200.

In one embodiment, the anti-microbial active group of this invention maybe selected from: (a) a tertiary amine (i.e. R₃ and/or R₃′ is nothing)or tertiary ammonium (i.e. R₃ and/or R₃′ is hydrogen) comprising atleast one terpenoid moiety (b) a quaternary ammonium group comprising atleast one terpenoid moiety (c) a quaternary ammonium group, comprisingat least one alkyl group having from 4 to 24 carbon atoms; and (d) atertiary amine (i.e. R₃ and/or R₃′ is nothing) or tertiary ammonium(i.e. R₃ and/or R₃′ is hydrogen) comprising at least one alkyl grouphaving from 4 to 24 carbon atoms. Each possibility represents a separateembodiment of this invention.

In one embodiment, the particles of this invention represented bystructures (1)-(6) comprise an anti-microbial active unit and an inertcore, wherein the anti-microbial active unit and the core are linkedindirectly.

In some embodiments L₁, L₂ or L₃ is each independently the same or adifferent linker. In some embodiments, L₁, L₂ and L₃ are connected toeach other, in any possible way. In some embodiment, L₃ is nothing andL₁ or L₂ is connected to the core covalently. In another embodiment, L₃is connected to the core covalently and L₁ or L₂ is connected to L₃. Inanother embodiment, L₁ is connected to X, X′ and L₃ or core. In anotherembodiment, a “linker” comprises any possible chemical moiety capable ofconnecting at least two other chemical moieties which are adjacent tosuch linker. In another embodiment, the monomeric unit of theanti-microbial active unit comprises a first and/or second linker/s (L₁or L₂) and an anti-microbial group. In another embodiment, L₁ and/or L₂are/is the backbone of the anti-microbial active unit. In someembodiments, the linker comprises a functional group. In anotherembodiment, the linker comprises two (same or different) functionalgroups. In another embodiment, the functional group comprises phosphate,phosphonate, siloxane, silane, ether acetal, amide, amine, anhydride,ester, ketone, or aromatic ring or rings functionalized with any of thepreceding moieties. Each possibility represents a separate embodiment ofthis invention.

In another embodiment, L₁ or L₂ is a C1 to C18 alkylene, alkenylene,alkynylene or aryl substituted with at least one carboxyl moiety,wherein the carboxyl end is attached to the core. This linker may bederived from a C1 to C18 alkylene substituted with at least one carboxylmoiety and having an amino end which is modified to anti-microbiialactive group [—⁺N(R₁)(R₂)(R₃), —⁺NH(R₁)(R₂),—⁺N(R₁)(R₂)—⁺N(R₁′)(R₂′)(R₃′), —⁺NH(R₁′)(R₂′) or —N(R₁′)(R₂′) (definedin structures (1) to (6))]. This linker may be derived from an aminoacid of natural or synthetic source having a chain length of between 2and 18 carbon atoms (polypeptide), or an acyl halide of said amino acid.Non-limiting examples for such amino acids are 18-amino octadecanoicacid and 18-amino stearic acid. In another embodiment, L₁ or L₂ is a C1to C18 alkylene substituted with at least one amine or amide moiety.

In another embodiment, L₁, L₂, L₃ or any combination thereof is a C1 toC18 alkylene, alkenylene, alkynylene or aryl. This linker may be derivedfrom a di-halo alkylene, which is functionalized at each end with thecore and anti-microbial active group, respectively, by replacement ofthe halogen moiety to a functional group that binds to the core andreplacement of the halogen moiety to obtain [—⁺N(R₁)(R₂)(R₃),—⁺NH(R₁)(R₂), —N(R₁)(R₂)—⁺N(R₁′)(R₂′)(R₃′), —⁺NH(R₁′)(R₂′) or—N(R₁′)(R₂′) (defined in structures (1) to (6))]

In another embodiment, L₁, L₂, L₃ or any combination thereof is anaromatic group derived from non-limiting examples of 4,4-biphenol,dibenzoic acid, dibenzoic halides, dibenzoic sulphonates, terephthalicacid, tetrphthalic halides, and terephthalic sulphonates. This linker isfunctionalized with the core and anti-microbial active group,respectively, through the functional group thereof (i.e., hydroxyl,carboxy or sulfonate). In another embodiment, this linker is directlyattached to the core at one end or indirectly, via a third linker (L₃)and is modified at the other end to anti-microbial active group[—⁺N(R¹)(R₂)(R₃), —⁺NH(R₁)(R₂), —N(R₁)(R₂)—⁺N(R₁′)(R₂′)(R₃′),—⁺NH(R₁′)(R₂′) or —N(R₁′)(R₂′) (defined in structures (1) to (6))].

In another embodiment, L₁, L₂, L₃ or any combination thereof, is asiloxane or silane group derived and/or selected from non-limitingexamples of trialkoxyalkylsilane, trialkoxyarylsilane,trihaloalkylsilane, trihaloarylsilane, 3-aminopropyltriethoxysilane(APTES) and N-2-aminoethyl-3-aminopropyl trimethoxysilane. This linkeris functionalized with the core and anti-microbial active group,respectively, through the functional group thereof (i.e., hydroxyl,siloxane, carboxy, amide or sulfonate). In another embodiment, thislinker is directly attached to the core at one end directly orindirectly, via a third linker (L3) and is modified at the other end toanti-microbial active group [—⁺N(R₁)(R₂)(R₃), —⁺NH(R₁)(R₂),—N(R₁)(R₂)—⁺N(R₁′)(R₂′)(R₃′), —⁺NH(R₁′)(R₂′) or —N(R₁′)(R₂′) (defined instructures (1) to (6))].

In one embodiment, the anti-microbial active group of this invention maybe selected from: (a) a tertiary amine (i.e. R₃ and/or R₃′ is nothing)or tertiary ammonium (i.e. R₃ and/or R₃′ is hydrogen) comprising atleast one terpenoid moiety (b) a quaternary ammonium group comprising atleast one terpenoid moiety (c) a quaternary ammonium group, comprisingat least one alkyl group having from 4 to 24 carbon atoms; and (d) atertiary amine (i.e. R₃ and/or R₃′ is nothing) or tertiary ammonium(i.e. R₃ and/or R₃′ is hydrogen) comprising at least one alkyl grouphaving from 4 to 24 carbon atoms. Each possibility represents a separateembodiment of this invention.

This linker is functionalized with the core and anti-microbial activegroup, respectively, through the functional group thereof (i.e.,hydroxyl, siloxane, carboxy, amide or sulfonate). In another embodiment,this linker is directly attached to the core at one end or indirectly,via a third linker (L3) and is modified at the other end toanti-microbial active group [—⁺N(R₁)(R₂)(R₃), —⁺NH(R₁)(R₂),—N(R₁)(R₂)—⁺N(R₁′)(R₂′)(R₃′), —⁺NH(R₁′)(R₂′) or —N(R₁′)(R₂′) (defined instructures (1) to (6))].

In another embodiment, a monomeric unit within the anti-microbial activeunit of this invention is represented by the structure of formula IA:

whereinR₁ and R₂ are independently linear or branched alkyl, terpenoid,cycloalkyl, aryl, heteroaryl, alkenyl, alkynyl or any combinationthereof, andR₃ is nothing, linear or branched alkyl, terpenoid, cycloalkyl, aryl,heteroaryl, alkenyl, alkynyl or any combination thereof, wherein if R₃is nothing, the nitrogen is not chargedq is an integer between 0 and 16;wherein said monomeric unit is chemically bound to the surface of aninorganic core directly or via a third linker (L3).

In another embodiment, a monomeric unit within the anti-microbial activeunit of this invention is represented by the structure of formula IB:

whereinR₁ and R₂ are independently linear or branched alkyl, terpenoid,cycloalkyl, aryl, heteroarylalkenyl, alkynyl or any combination thereof,andR₃ is nothing, linear or branched alkyl, terpenoid, cycloalkyl, aryl,heteroaryl, alkenyl, alkynyl or any combination thereof, wherein if R₃in nothing, the nitrogen is not chargedq and q¹ are independently an integer between 0 and 16;wherein said monomeric unit is chemically bound to the surface of aninorganic core directly or via a third linker (L3).

In another embodiment, a linker molecule within the anti-microbialactive unit of this invention is represented by the structure of formulaIC:

whereinQ²⁰¹, Q²⁰² and Q²⁰³ are independently selected from the group consistingof alkoxy, methyl, ethyl, hydrogen, sulfonate and halide, wherein atleast one of Q²⁰¹, Q²⁰² and Q²⁰³ is selected from ethoxy, methoxy,sulfonate (e.g., mesyl, tosyl) and halide; andq is an integer between 0 and 16;the linker molecule is capable of being chemically bound to the surfaceof the inorganic core through the silicon atom; andthe anti-microbial active group is introduced by functionalizing theprimary amine to obtain an anti-microbial active tertiary amine orquaternary ammonium group containing at least one terpenoid group, asdescribed above.

In another embodiment, a linker molecule within the anti-microbialactive unit of this invention is represented by the structure of formulaID:

whereinQ²⁰¹, Q²⁰² and Q²⁰³ are independently selected from the group consistingof alkoxy, methyl, ethyl, hydrogen, sulfonate and halide, wherein atleast one of Q²⁰¹, Q²⁰² and Q²⁰³ is selected from ethoxy, methoxy,sulfonate (e.g., mesyl, tosyl) and halide;W is selected from the group consisting of NH₂, halide, sulfonate andhydroxyl; andq is an integer between 0 and 16;said linker is capable of being chemically bound to the surface of saidinorganic core through the silicon atom; andthe anti-microbial active group is introduced by substituting the groupW with an anti-microbial active group, or converting the group W to ananti-microbial active group.

The particles of this invention demonstrate an enhanced anti-microbialactivity. Without being bound by any theory or mechanism, it can bepostulated that such activity originates from the presence of closelypacked anti-microbial groups on a given core's surface, as well as highdensity of particles packed on the surface of a host material. Thisdensity increases as each anti-microbial active unit in the particles ofthis invention comprise increasing number of anti-microbial activegroups and it yields a high local concentration of active functionalgroups, which results in high effective concentration of theanti-microbial active groups and enables the use of a relatively smallamount of particles to achieve effective bacterial annihilation. Theclose packing of the anti-microbial groups is due to, inter alia,numerous anti-microbial active units protruding from each particlesurface. Accordingly, the anti-microbial groups cover large fraction ofthe particle's available surface area (width dimension covering thesurface). The surface density of the anti-microbial group results inhigh effective concentration promoting anti-microbial inhibitory effect.According to the principles of this invention, high surface densitydictates high anti-microbial efficiency.

The term “nanoparticle” as used herein refers to a particle having adiameter of less than about 1,000 nm. The term “microparticle” as usedherein refers to a particle having a diameter of about 1,000 nm orlarger.

The anti-microbial particles of this invention are characterized byhaving a diameter between about 5 to about 100,000 nm, and thusencompass both nanoparticulate and microparticulate compositions.Preferred are particles between about 10 to about 50,000 nm. In otherembodiments, the particles are more than 1,000 nm in diameter. In otherembodiments, the particles are more than 10,000 nm in diameter. In otherembodiment, the particles are between 1,000 and 50,000 nm in diameter.In other embodiment, the particles are between 5 and 250 nm in diameter.In other embodiment, the particles are between 5 and 500 nm in diameter.In another embodiment, the particles are between 5 and 1000 nm indiameter. It is apparent to a person of skill in the art that otherparticles size ranges are applicable and are encompassed within thescope of this invention.

Anti-Microbial Active Groups Comprising Terpenoid Groups

In one embodiment, the anti-microbial active group of this inventioncontains at least one terpenoid group, and is selected from: (a) atertiary amine (R₃ and/or R₃′ is nothing) or tertiary ammonium (R₃and/or R₃′ is H) comprising at least one terpenoid moiety; and (b) aquaternary ammonium group comprising at least one terpenoid moiety.

In some embodiments, the anti-microbial active group of formula (1) to(6) is selected from: (a) a tertiary amine (R₃ and/or R₃′ is nothing) ortertiary ammonium (R₃ and/or R₃′ is H), wherein the nitrogen atom ofeach tertiary amine/ammonium having at least one bond to X₁ or X₂ andone bond to a terpenoid moiety; (b) a tertiary amine (R₃ and/or R₃′ isnothing), or tertiary ammonium (R₃ and/or R₃′ is H), the nitrogen atomof each tertiary amine/ammonium having one bond to X₁ or X₂ and twobonds to terpenoid moieties which may be the same or different from eachother, or a salt of said tertiary amine; (c) a quaternary ammonium groupthe nitrogen atom of each quaternary ammonium group having at least onebond to X₁ or X₂ and one or two bonds to terpenoid moieties which may bethe same or different from each other; Each possibility represents aseparate embodiment of this invention.

The term “terpenoid”, also known as “isoprenoid” refers to a large classof naturally occurring compounds that are derived from five-carbonisoprene units.

In one embodiment, the at least one terpenoid moiety is a cinammyl groupderived from cinnamaldehyde, cinnamic acid, curcumin, viscidone orcinnamyl alcohol. In another embodiment, the at least one terpenoidmoiety is a bornyl group derived from camphor, bornyl halide or bornylalcohol. In another embodiment, the at least one terpenoid moiety isderived from citral. In another embodiment, the at least one terpenoidmoiety is derived from perilaldehyde. Each possibility represents aseparate embodiment of this invention.

Cinnamaldehyde is a natural aldehyde extracted from the genusCinnamomum. It is known for its low toxicity and its effectivenessagainst various bacteria and fungi.

Camphor is found in the wood of the camphor laurel (Cinnamomumcamphora), and also of the kapur tree. It also occurs in some otherrelated trees in the laurel family, for example Ocotea usambarensis, aswell as other natural sources. Camphor can also be syntheticallyproduced from oil of turpentine. Camphor can be found as an R or Senantiomer, a mixture of enantiomers and a racemic mixture. Eachpossibility represents a separate embodiment of this invention.

Citral, or 3,7-dimethyl-2,6-octadienal or lemonal, is a mixture of twodiastereomeric terpenoids. The two compounds are double bond isomers.The E-isomer is known as geranial or citral A. The Z-isomer is known asneral or citral B. Citral is known to have anti-microbial activity.

Perillaldehyde, also known as perilla aldehyde, is a natural terpenoidfound most in the annual herb perilla, as well as in a wide variety ofother plants and essential oils.

Other examples of terpenoids include, but are not limited to:curcuminoids found in turmeric and mustard seed, citronellal found inCymbopogon (lemon grass) and carvacrol, found in Origanum vulgare(oregano), thyme, pepperwort, wild bergamot and Lippia graveolens. Eachpossibility represents a separate embodiment of this invention.

In accordance with the above embodiment, the anti-microbial activeterpenoid moieties are selected from the group consisting of:

or any combination thereof,

Each possibility represents a separate embodiment of this invention.

Non-limiting examples of functional anti-microbial active tertiary aminegroups or its protonated form in accordance with the principles of thisinvention are:

wherein R² is alkyl, terpenoid, cycloalkyl, aryl, heterocycle, alkenyl,alkynyl or any combination thereof.

Non-limiting examples of anti-microbial active quaternary ammoniumgroups in accordance with the principles of this invention are:

wherein R² is alkyl, terpenoid, cycloalkyl, aryl, heterocycle, alkenyl,alkynyl or any combination thereof;R³ is alkyl, terpenoid moiety, cycloalkyl, aryl, heterocycle, alkenyl,alkynyl or any combination thereof;

The anti-microbial active group of this invention may be in the form ofa tertiary amine, or in the form of a protonated said tertiary amine, orin the form of a quaternary ammonium salt, as described hereinabove.Since an ammonium group is positively charged, its charge is balancedwith an anion. Preferably, in a particle according to this inventionthis anion is a halide, e.g. fluoride, chloride, bromide or iodide, andfluoride is most preferred. Other possible anions include, but are notlimited to, bicarbonate, nitrate, phosphate, acetate, fumarate,succinate and sulfate. Each possibility represents a separate embodimentof this invention.

Anti-Microbial Active Groups Comprising One Long Alkyl Group.

In accordance with another embodiment, the anti-microbial active groupof this invention [—⁺N(R₁)(R₂)(R₃), —⁺NH(R₁)(R₂),—N(R₁)(R₂)—⁺N(R₁′)(R₂′)(R₃′), —⁺NH(R₁′)(R₂′) or —N(R₁′)(R₂′) (defined instructures (1) to (6))] is a quaternary ammonium group, a tertiary amineor a tertiary ammonium, the nitrogen atom of each amine/ammonium grouphaving at least one bond X1 or X₂, at least one bond to an alkyl grouphaving from 4 to 24 carbon atoms (R₁ and/or R₁′). In another embodiment,the nitrogen atom of each amine/ammonium group having one bond to thecore, one bond to an alkyl group having from 4 to 24 carbon atoms (R₁and/or R₁′).

Since an ammonium group is positively charged, its charge should bebalanced with an anion. Any of the counter-ions described above may beused to counter-balance the quaternary ammonium group.

In some embodiments, the nitrogen atom of each quaternary ammonium ortertiary ammonium group has (i) at least one bond to X₁ or X₂; and (ii)at least one bond to the alkyl group having from 4 to 24 carbon atoms.

In some embodiments, the anti-microbial active group of formula (1) to(6) is selected from: (a) a tertiary amine (R₃ and/or R₃′ is nothing) ortertiary ammonium (R₃ and/or R₃′ is H), wherein the nitrogen atom ofeach tertiary amine/ammonium having at least one bond to X₁ or X₂ andone bond to the alkyl group having from 4 to 24 carbon atoms; (b) atertiary amine (R₃ and/or R₃′ is nothing), or tertiary ammonium (R₃and/or R₃′ is H), wherein the nitrogen atom of each tertiaryamine/ammonium having one bond to X₁ or X₂ and two bonds to alkyl groupshaving from 4 to 24 carbon atoms which may be the same or different fromeach other, or a salt of said tertiary amine; (c) a quaternary ammoniumgroup wherein the nitrogen atom of each quaternary ammonium group havingat least one bond to X₁ or X₂ and one or two bonds to the alkyl groupshaving from 4 to 24 carbon atoms which may be the same or different fromeach other. Each possibility represents a separate embodiment of thisinvention.

The term “quaternary ammonium group” refers to a group of atomsconsisting of a nitrogen atom with four substituents (different thanhydrogen) attached thereto. In another embodiment, a “quaternaryammonium group” refers to a group of atoms consisting of a nitrogen atomwith four groups wherein each of the group is attached to the nitrogenthrough a carbon atom. The term “long alkyl group” or chain refers tosuch an alkyl group or chain which is substituted on the nitrogen atomof the quaternary ammonium group and which has between 4 and 24 carbonatoms. In some currently preferred embodiments, the alkyl group is analkyl group having 4 to 18 carbon atoms. In some currently preferredembodiments, the alkyl group is an alkyl group having 4 to 8 carbonatoms. In some currently preferred embodiments, the alkyl group is analkyl group having 4 to 10 carbon atoms. In other currently preferredembodiments, the alkyl group is an alkyl group having 6, 7, or 8 carbonatoms, with each possibility representing a separate embodiment of thisinvention.

Organic Polymeric Cores

In some embodiments, the core of the anti-microbial particles is anorganic polymeric core. In one embodiment, the organic core comprises atleast one aliphatic polymer. An “aliphatic polymer” as used within thescope of this invention refers to a polymer made of aliphatic monomersthat may be substituted with various side groups, including (but notrestricted to) aromatic side groups. Aliphatic polymers that may beincluded in particles according to this invention comprise nitrogenatoms (as well as other heteroatoms) as part of the polymeric backbone.In one embodiment, the core of the particles is an organic polymericcore including an amine which can be substituted with R₁, R₂ and/or R₃as defined for structure 1; or including an imine which is chemicallymodified to amine and then substituted with R₁, R₂ and/or R₃ as definedfor structure 1. Non-limiting examples of aliphatic polymers arepolystyrene (PS), polyvinylchloride (PVC), polyethylene imine (PEI),polyvinyl amine (PVA), poly(allyl amine) (PAA), poly(aminoethylacrylate), polypeptides with pending alkyl-amino groups, and chitosan.Each possibility represents a separate embodiment of this invention. Inone currently preferred embodiment, the polymer is polyethylene imine(PEI).

In another embodiment, the organic core comprises at least one aromaticpolymer selected from the following group: polystyrene, aminomethylatedstyrene polymers, aromatic polyesters, preferably polyethyleneterephthalate, and polyvinyl pyridine.

The polymeric core may be linked to anti-microbial active part directly(i.e. in structures (1)-(3): L₃ is a bond) or via a linker. Eachpossibility represents a separate embodiment of this invention.

In one embodiment, the organic polymeric core includes a combination oftwo or more different organic polymers. In another embodiment, theorganic polymeric core includes a copolymer.

In some embodiments, anti-microbial active unit is linked to the organicpolymeric core directly (L₃ is a bond) or via a linker (L₃). In theseembodiments, the linker may be selected from:

(a) a C1 to C18 alkylene substituted with at least one carboxyl moiety.This linker may be derived from an alkylene substituted with at leastone carboxyl moiety and at least one amino moiety, wherein the carboxylend is attached to the core and the amino end is modified toanti-microbial active group [—⁺N(R₁)(R₂)(R₃), —⁺NH(R₁)(R₂),—N(R₁)(R₂)—⁺N(R₁′)(R₂′)(R₃′), —⁺NH(R₁′)(R₂′) or —N(R₁′)(R₂′) (defined instructures (1) to (6))]. This linker may be derived from an amino acidof natural or synthetic source having a chain length of between 2 and 18carbon atoms, or an acyl halide of said amino acid. Non-limitingexamples for such amino acids are 18-amino octadecanoic acid and18-amino stearic acid;(b) a C1 to C18 alkylene. This linker may be derived from a di-haloalkylene, which is functionalized at each end with the core andanti-microbial active group, respectively, by replacement of the halogenmoiety to a functional group that will bind to the core and replacementof the halogen moiety to obtain [—⁺N(R₁)(R₂)(R₃), —⁺NH(R₁)(R₂),—N(R₁)(R₂)—⁺N(R₁′)(R₂′)(R₃), —⁺NH(R₁′)(R₂′) or —N(R₁′)(R₂′) (defined instructures (1) to (6))]; and (c) aromatic molecules derived from4,4-biphenol, dibenzoic acid, dibenzoic halides, dibenzoic sulphonates,terephthalic acid, tetrphthalic halides, and terephthalic sulphonates.This linker is functionalized with the core and anti-microbial activegroup, respectively, through the functional group thereof (i.e.,hydroxyl, carboxy or sulfonate). In another embodiment, this linker isattached to the core at one end and is modified at the other end toanti-microbial active group [—⁺N(R₁)(R₂)(R₃), —⁺NH(R₁)(R₂),—N(R₁)(R₂)—⁺N(R₁′)(R₂′)(R₃′), —⁺NH(R₁′)(R₂′) or —N(R₁′)(R₂′) (defined instructures (1) to (6))] In another embodiment, the linker comprisesalkyl, alkenyl, alkyl phosphate, alkyl siloxanes, carboxylate, epoxy,acylhalides and anhydrides, or combination thereof, wherein thefunctional group is attached to the core. Each possibility represents aseparate embodiment of this invention.

Various polymeric chains may provide a range of properties thatthemselves may be an accumulation of the various polymer properties, andmay even provide unexpected synergistic properties. Examples of suchmixed polyamine particles include: crosslinking of aliphatic andaromatic polyamines such as polyethyleneimine and poly(4-vinyl pyridine)via a dihaloalkane; mixture of linear short chain and branched highmolecular weight polyethyleneimines; interpenetrating compositions ofpolyamine within a polyamine scaffold such as polyethyleneimine embeddedwithin crosslinked polyvinyl pyridine particles, or eveninterpenetrating a polyamine into a low density non-amine scaffold suchas polystyrene particles. In other words, the use of polyaminecombinations for the purpose of forming particles, either by chemicalcrosslinking or physical crosslinking (interpenetrating networks) mayafford structures of varying properties (such as being able to betterkill one bacteria vs. another type of bacteria). Such properties may beadditive or synergistic in nature.

In one specific embodiment, the organic polymeric core is cross-linkedwith a cross-linking agent. The preferred degree of cross-linking isfrom 1% to 20%, when crosslinking of from about 2% to about 5% ispreferable. The crosslinking may prevent unfolding of the polymer andseparation of the various polymeric chains that form the particle.

Crosslinking, as may be known to a person skilled in the art of organicsynthesis and polymer science, may be affected by various agents andreactions that are per se known in the art. For example, crosslinkingmay be affected by alkylating the polymer chains with dihaloalkane suchas dibromoethane, dibromocyclohexane, or bis-bromomethylbenzene.Alternatively, crosslinking by reductive amination may be used. In thismethod a polyamine with primary amines is reacted with a diketone orwith an alkane dialdehyde to form an imine crosslinker which is thenfurther hydrogenated to the corresponding amine. This amine is furtherreacted to form an antimicrobial effective quaternary ammonium group. Insuch a method, instead of dihaloalkanes or dialdehydes, tri orpolyhaloalkanes or polyaldehydes or polyketones are used.

The preferred polymers useful for making the polymeric core according tothis invention are those having chains made of 30 monomer units,preferably 100 monomer units that may be crosslinked using less than 10%of crosslinking agent. The longer the polymers are, the fewercrosslinking bonds are needed to afford an insoluble core. Branchedpolymers are preferred for crosslinking as small amount of crosslinkingis required to form insoluble core.

In some embodiments, at least about 10% of the amine groups in theorganic polymeric core are the anti-microbial active tertiaryamine/ammonium or quaternary ammonium groups or salts thereof, asdescribed herein.

In one embodiment, the anti-microbial particles according to thisinvention have functional groups that are capable of reacting with ahost polymer or with monomers thereof. Such functional groups aredesigned to allow the particles to be bound chemically to a hostingmaterial.

Inorganic Cores

In some embodiments, the core of the anti-microbial particles of thisinvention is an inorganic core comprising one or more inorganicmaterials. Inorganic cores have a few advantages over organic polymericcores: 1) higher stability at elevated temperature; 2) higher chemicalstability towards various solvent and reagents; 3) improved mechanicalstrength; 4) better handling qualities in composites due to theiramphipathic nature; and 5) lower cost.

An additional advantage of inorganic cores relates to the insertion ofthe functionalized particles into a polymeric material within apolymeric matrix (host). In contrast to organic cores which are producedby radical polymerization (e.g. acrylate resins), inorganic cores do notinterfere with the polymerization process and hence do not jeopardizethe mechanical properties of the finalized substrate, as opposed toorganic polymeric cores which tend to interfere with the polymerizationreaction.

In one embodiment, the inorganic core comprises silica, metal, metaloxide or a zeolite. Each possibility represents a separate embodiment ofthis invention.

In one embodiment, the core of the particles of this invention comprisessilica (SiO₂). The silica may be in any form known in the art,non-limiting examples of which include polyhedral oligomericsilsesquioxane (POSS), amorphous silica, dense silica, aerogel silica,porous silica, mesoporous silica and fumed silica.

The surface density of active groups onto particle surface haveproportional impact on its anti-microbial activity. This is applicableboth to organic and inorganic particles in same manner. In anotherembodiment, the core of the particles of this invention comprisesglasses or ceramics of silicate (SiO₄ ⁻⁴). Non-limiting examples ofsilicates include aluminosilicate, borosilicate, barium silicate, bariumborosilicate and strontium borosilicate.

In another embodiment, the core of the particles of this inventioncomprises surface activated metals selected from the group of: silver,gold, platinum, palladium, copper, zinc and iron.

In another embodiment, the core of the particles of this inventioncomprises metal oxides selected from the group of: zirconium dioxide,titanium dioxide, vanadium dioxide, zinc oxide, copper oxide andmagnetite.

The inorganic core typically has a solid uniform morphology with lowporosity or a porous morphology having pore size diameter of betweenabout 1 to about 30 nm.

In another embodiment, the core of the particles of this inventioncomprises natural or artificial Zeolites.

In one embodiment, the core may be attached to the anti-microbial unitdirectly (i.e. in structures (1)-(3): L₃ is a bond), or via a linker(L₃). Preferably a silica (SiO₂) based inorganic core may be attached tothe anti-microbial part through a linker (L₃), while glasses orceramicas of silicate (SiO₄ ⁻⁴), metals or metal oxides may be attachedto anti-microbial unit directly (i.e. in structures (1)-(3): L₃ is abond).

In some embodiments, the inorganic core is directly (i.e. in structures(1)-(3): L₃ is a bond) attached to the anti-microbial unit. In otherembodiments, the inorganic core is attached to the anti-microbial unitthrough a linker. In some embodiments, the linker is selected from thefollowing groups: a C1 to C18 alkylene; a C1 to C18 alkylene substitutedwith at least one silane or alkoxysliane moiety; a C1 to C18 alkylenesubstituted with at least one phosphate moiety; a C1 to C18 alkylenesubstituted with at least one anhydride moiety; a C1 to C18 alkylenesubstituted with at least one carboxylate moiety; and a C1 to C18alkylene substituted with at least one glycidyl moiety. Each possibilityrepresents a separate embodiment of this invention.

The inorganic core of the particle as described above may generally bein a form selected from a sphere, amorphous polygonal, shallowflake-like and a rod. In some representative embodiments, the inorganiccore is spherical and has a diameter between about 5 to about 100,000nm. In some representative embodiments, the inorganic core is sphericaland has a diameter between about 1000-100,000 nm. In some representativeembodiments, the inorganic core is spherical and has a diameter betweenabout 100-1000 nm with pore diameter of about 1 to about 100 nm. Inanother embodiment, the inorganic spherical core has a pore diameter ofabout 1 to about 50 nm. In another embodiment, the inorganic sphericalcore has a pore diameter of about 1 to about 30 nm. In anotherembodiment, the inorganic particle is in a form of a rod, having adiameter of between about 5 to about 1,000 nm and length between about10 to about 1,000,000 nm. In another embodiment, a length of between 50to 100,000 nm. In another embodiment, a length of between 100 to 250,000nm. In another embodiment, a length of between 200 to 500,000 and a porediameter of about 1 to about 50 nm. Each possibility represents aseparate embodiment of this invention.

Preparation of Anti-Microbial Particles, Comprising One Monomeric UnitPer One Anti-Microbial Active Part

The particles of this invention may be prepared in accordance to avariety of processes, depending on the nature of the core, theanti-microbial active group, and the presence or absence of linkers.Some non-limiting examples of preparation methods are provided below.

In one embodiment, this invention provides processes for preparinganti-microbial particles, wherein the particles comprise one monomericunit per one anti-microbial active unit. In the following, suchprocesses will be presented in detail.

A representative method for preparing particles according to thisinvention wherein the anti-microbial active group is a tertiary amine ora quaternary ammonium group comprising at least one terpenoid moiety isrepresented in FIG. 2. In accordance with FIG. 2, a core as definedherein is functionalized with a primary amine. The primary amine reactswith an aldehyde to yield initially an imine (Schiff base) intermediateof formula (A′), which is then reacted with a second aldehyde underreductive amination conditions to yield a tertiary amine of formula(B′). RC(═O)H and R′C(═O)H each represent an aldehyde which is aterpenoid or which is derived from a terpenoid. RC(═O)H and R′C(═O)H maybe the same or different from each other. Conversion of the tertiaryamine to the quaternary ammonium group is optional, and involvesreaction of the tertiary amine with a group R¹—Y wherein R¹ is a C₁-C₄alkyl and Y is a leaving group such as halogen or sulfonate.

It is understood that the group

may represents any one or more of the following:1. An organic core directly bound to NH₂.2. An organic core bound to NH₂ through a linker as described herein.3. An inorganic core directly bound to NH₂.4. An inorganic core bound to NH₂ through a linker as described herein.

The exemplified reaction (FIG. 2) may be a “one pot synthesis”, or itmay include two sequential reactions with isolation of an intermediateformed in the first step. The first step is the formation ofintermediate (A′), which is an imine (Schiff base), by reacting an aminefunctionalized core with a terpenoid moiety in the presence of areducing agent, in this case cinnamyl in the presence of NaBH₄. Theimine functionalized core can be isolated at this stage if desired.Alternatively, further reacting intermediate (A′) with a terpenoidmoiety in the presence of a reducing agent yields a tertiary aminecomprising two terpenoid moieties (B′). In order to obtain thequaternary ammonium, additional alkylation step is performed asdescribed in FIG. 2.

The process presented in FIG. 3 uses cinnamaldehyde, but is applicableto other aldehydes. Thus, in some embodiments, this invention provides aparticle comprising (i) an inorganic core or an organic polymeric core;and (ii) an imine moiety chemically bound to the core, preferably at asurface density of at least one imine group per 10 sq. nm, wherein theimine group comprises a terpenoid moiety. The imine moiety is generallyrepresented by the structure of formula (B′) in FIG. 2. A more specificembodiment is the structure of formula (B) in FIG. 3. It is understoodby a person of skill in the art that other imine intermediate compoundscomprising other terpenoids groups as described herein, are alsoencompassed by this invention.

A representative method for preparing particles according to thisinvention wherein the anti-microbial active group is a quaternaryammonium group containing one alkyl group having 4 to 18 carbon atoms ispresented in FIG. 4. The method includes three pathways to preparequaternary ammonium salts (QAS) functionalized particle. A) by firstutilizing reductive amination to achieve tertiary amine, followed by analkylation reaction, B) by stepwise alkylation reactions; and C) byreacting a linker functionalized with a leaving group (e.g., Cl or otherhalogen) with tertiary amine. R¹ and R² represent C₁-C₄ alkyls such asmethyl, ethyl, propyl or isopropyl. R¹ and R² may be different or thesame group. Y represents any leaving group, for example Cl, Br or I, ora sulfonate (e.g., mesyl, tosyl).

It is understood that that the group

has any one of the meanings as described above for FIGS. 2 and 3.

It is understood that that the group

may represents any one or more of the following:1. An organic core directly bound to Y.2. An organic core bound to Y through a linker as described herein.3. An inorganic core directly bound to Y.4. An inorganic core bound to Y through a linker as described herein.

Core functionalization can occur by a solid support method, or asolution method (FIGS. 2-6).

Solid Support as Method of Preparation of Anti-Microbial ParticlesComprising One Monomeric Unit Per One Anti-Microbial Active Part

Preparation of functionalized particles is conducted in two generalsteps. First, the linker molecule is allowed to condense onto particlessurface (surface functionalization) via hydrolysis of leaving groups togive an intermediate of formula (FIG. 5, D′). Second, functional sitesof the linker molecule undergo further functionalization (linkerfunctionalization) as mentioned in any ones of (FIGS. 2-4) to give afunctionalized particle of formula (E′).

Solution Method as Method of Preparation of Anti-Microbial ParticlesComprising One Monomeric Unit Per One Anti-Microbial Active Part

In this method, the linker molecule is first functionalized withantimicrobial active group to give an intermediate of formula (FIG. 5,F′). In the second stage intermediate (F′) is allowed to settle ontoparticle's solid surface (surface functionalization) to give afunctionalized particle of formula (FIG. 5, E′).

This process is exemplified in FIG. 6 for cinnamaldehyde, but isapplicable to other aldehydes.

Preparation of Anti-Microbial Particles, Comprising More than OneMonomeric Unit Per One Anti-Microbial Active Unit

In one embodiment, this invention provides processes for preparingparticles of the composites of this invention, wherein the particlescomprise more than one monomeric unit per one anti-microbial activeunit. In the following, such processes will be presented in detail.

Solid Support as Method of Preparation of Anti-Microbial ParticlesComprising More than One Monomeric Unit Per One Anti-Microbial ActiveUnit

The solid support method comprises a few stages. First, the linkermolecule (dilute solutions of a few percent) is allowed to condense ontoparticles surface (surface functionalization) via (acid catalyzed)hydrolysis of leaving groups, resulting in the attachment of the linkerto the core (FIG. 7, step 1). Second, the attached linker is elongated.In another embodiment, this stage is achieved synthetically via one stepor more. In another embodiment, elongation is achieved by consecutiveaddition of difunctionalized alkane and diaminoalkane, wherein amines(of attached linker and diaminoalkane) attack electrophilic centers ofthe difunctionalized alkane (FIG. 7, steps 2 and 3). In anotherembodiment, such consecutive addition is optionally repeated for 1-10times. Finally, the anti-microbial active group (usually attached to analkylene chain) is grafted to resulting attached and elongated linker.In another embodiment, grafting is accomplished when amines on theattached and elongated linker attack acyl halide moiety of the moleculeof the anti-microbial active group which is grafted (FIG. 7, step 4).

In another embodiment, the same trialkoxysilane linker molecule is usedinitially, however in a higher concentration (≥10% by wt) and itinitially self-polymerizes (FIG. 8A) under basic catalysis.Functionalization of the solid supported linker progresses similarly asin the procedures described hereinabove for particles that comprise moreone monomeric unit per one anti-microbial active unit (FIGS. 2-5).

Solution Method as Method of Preparation of Anti-Microbial ParticlesComprising More than One Monomeric Unit Per One Anti-Microbial ActiveUnit

The solution method comprises a few stages. The first step involveselongation of the linker molecule. In another embodiment, this step isachieved synthetically via one step or more. In another embodiment,elongation is achieved by consecutive addition of difunctionalizedalkane and diaminoalkane wherein amines (of linker and diaminoalkane)attack electrophilic centers of the difunctionalized alkane (FIG. 9,steps 1 and 2). In another embodiment, such consecutive addition isoptionally repeated for 1-10 times. In the second stage, theanti-microbial active group (usually attached to an alkylene chain) isgrafted to resulting elongated linker. In another embodiment, graftingis accomplished when amines on the elongated linker attack acyl halidemoiety of the molecule of the anti-microbial active group which isgrafted (FIG. 9, step 3). Finally, the elongated, anti-microbial activelinker is attached to the core via functionalization thereof. In thisstep, the linker molecule (dilute solutions of a few percent) is allowedto condense onto particles surface (surface functionalization) via (acidcatalyzed) hydrolysis of leaving groups, resulting in the attachment ofthe linker to the core (FIG. 9, step 4).

This process is exemplified in FIGS. 10-11 for silica functionalizedwith dimethylethylammonium, but is applicable to otherhydroxyl-terminated cores and anti-microbial active groups.

In another embodiment, the same trialkoxysilane linker molecule is usedinitially, however in a higher concentration (≥10% by weight) and itinitially self-polymerizes (FIG. 8B) under basic catalysis.Functionalization of the linker progresses similarly as in theprocedures described hereinabove for particles that comprise more onemonomeric unit per one anti-microbial active part (FIGS. 2-5).

Preparation of Core Particles

In some embodiments, the particles of the composites of this inventionwhich comprise one or more monomeric units per one anti-microbial activepart, comprise cores which are prepared according to the following.

Porous silica materials can be prepared by reaction of SiCl₄ withalcohol or water, followed by drying using centrifugation and/or heatingutilizing airflow or under vacuum conditions. Dense fumed silicaparticles (pyrogenic) were prepared by pyrolysis of SiCl₄.

An alternative preparation method of silica core material can be carriedby the hydrolysis of tetraethylorthosilicate (TEOS) or tetramethylorthosilicate (TMS) in the presence of alcohol or water solution andunder basic (Stober) or acidic catalytic conditions.

Mesoporous silica particles can be prepared by hydrolysis of TEOS or TMSat low temperatures, preferably in a temperature not exceeding 60° C.,followed by dehydration by centrifugation and/or evaporation underairflow or vacuum conditions.

Dense particles can be prepared utilizing intense heating in a processcalled calcination. Typically, such process takes place at hightemperatures at about 250° C.

Composition Comprising the Particles of this Invention

In some embodiments, the composition of this invention comprises theanti-microbial particles of this invention and a polymeric materialcomprising organic polymers, inorganic polymers or any combinationthereof. In some embodiment, the particles as described herein aredispersed in the polymeric material. In another embodiment, theparticles are homogeneously dispersed within the polymeric material. Inanother embodiment, the particles are found in the surface of thepolymeric materials. In another embodiment, the particles coat thepolymeric materials. In another embodiment, the particles interactweakly or physically (mechanically) with the polymeric material. Inanother embodiment, the anti-microbial particles are mechanicallyembedded within the polymeric material. In another embodiment, theseparticles are three dimensionally “locked” between the polymer chains,preventing them from migrating out from the complex network. The stronghydrophobic nature of these particles also plays a role in preventingthe particles from moving into the hydrophilic surrounds such as in thecase of physiological, dental, orthopedic or other medical applications.In another embodiment, the polymeric material is inert to the particlesand does not react with them. In one embodiment, the particles comprisefunctional groups, capable of reacting with moieties of the polymericmaterial. In another embodiment, the particles interact chemically withthe polymeric material. In another embodiment, the particles are amixture of different particles.

In some embodiments, the composition of this invention comprises theanti-microbial particles of this invention and a polymeric materialcomprising organic polymers, inorganic polymers or any combinationthereof. In another embodiment, the polymeric material comprisesthermoplastic polymers, thermoset polymers or any combination thereof.In another embodiment, the organic polymer comprises hydrogels,polyolefins such as polyvinylchloride (PVC), polyethylene, polystyreneand polypropylene, epoxy resins, acrylate resins such as poly methylmethacrylate, polyurethane or any combination thereof. In anotherembodiment, the inorganic polymer comprise silicone polymers such aspolydimethylsiloxane (PDMS), ceramics, metals or any combinationthereof. In another embodiment, the hydrogel is poloxamer or alginate.In another embodiment, the commercial poloxamer is used or it is formedby a reaction between a polymer and other reagent. In anotherembodiment, the polymer is poly(ethylene glycol) (PEG) with reactive endgroups (such as epoxides in PEG-diglycidyl ether) and the reagent hasmultiple reactive sites (e.g. diethylenetriamine). Each possibilityrepresents a separate embodiment of this invention.

In some embodiments, the weight ratio of the particles to the polymericmaterial is between 0.25-5%. In another embodiment, the weight ratio isbetween 0.5-2%. In another embodiment, the weight ratio is between 1-5%.

Another polymer material to be used in the context of this invention isresins used in dental, surgical, chirurgical and orthopedic compositematerials. In such applications, anti-microbial particles could be firstdispersed within the resin part or added simultaneously with filler orany other solid ingredients (if any). Most of these resins are acrylicor epoxy type monomers that undergo polymerization in-vivo.

Preparation of the Compositions of this Invention

In some embodiments, the composites of this invention are prepared byembedding the anti-microbial particles into the polymeric materials ofthis invention. In another embodiment, one type of particle is embeddedin the polymeric materials. In another embodiment, a combination ofdifferent particle types is embedded in the polymeric materials. In someembodiments, the embedding may be achieved by a variety ofmethodologies.

In some embodiments, embedding functionalized microparticles into apolymeric material is obtained by two methodologies: A) Extrusiontechnology: the particles are added into molten thermoplastic polymerinto extruder, preferably twin-coned extruder. B) A thermoplastic orthermoset polymer is heated in an organic solvent (non-limiting examplescomprise xylene, toluene, their derivatives or any combination thereof)under reflux conditions to achieve the complete dissolution of thepolymer. The anti-microbial particles are then dispersed in the samesolvent as used for the polymer and the mixture is added to thedissolved polymer using overhead stirrer or homogenizer. After completedispersion of particles within the polymer, the solvent is evaporatedusing conventional distillation or evaporation method.

In some embodiments, embedding functionalized microparticles into asilicone based polymeric material is obtained by several methodologies:A) Room temperature vulcanization (RTV) of silicone precursor isachieved, wherein particles are incorporated into unpolymerized orpre-polymerized silicone before final curing at final concentration of0.5-8% wt particles/silicone polymer. In another embodiment, the curingis activated by moisture. In another embodiment, the curing is activatedby heat. B) RTV of silicone precursor is achieved, whereinpolymerization is induced by mixing two components of the polymerizationmixture. In another embodiment, particles are incorporated into bothparts at final concentration of 0.5-8% wt. particles/silicone polymer,or in one of the parts at doubled concentration, giving the 0.5-8% wt.particles/silicone polymer final concentration.

Thus, according to some embodiments, this invention provides a methodfor preparing a composition comprising embedding a plurality ofanti-microbial particles in a polymeric material as described above,wherein the particles are embedded in the material, the method comprisesa step of adding the particles as described above, into a molten polymermaterial utilizing extrusion or to a polymer solution in solvent or viapolymerization with the particles and polymer precursors.

In some embodiments, particles according to this invention arehomogeneously distributed on the outer surface of the polymeric materialin a surface concentration of between about 0.1 to about 100 particlesper sq. micrometer. In another embodiment, particles according to thisinvention are homogeneously distributed on the outer surface of thepolymeric material in a surface concentration of between about 1 toabout 100 particles per sq. micrometer. The term “homogeneousdistribution” is used to denote a distribution, characterized in thatthe standard deviation of the number of particles per sq. um is no morethan the average number of particles per sq. micrometer. A homogeneousdistribution is preferred for reproducibility and productspecifications. If the distribution is not even, the product may exhibitdifferent properties at different areas. The distribution of theparticles away from the outer surface, that is, their bulkconcentration, may be similar to that on the outer surface. As a generalrule, the total surface of the particles preferably occupies at mostabout 20% of the surface of the material, preferably between 1% to 15%,more preferably between 1% and 5% and most about between 1% and 3% ofthe surface of the material.

According to some embodiments, on the average, every sq. micrometer ofthe outer surface of polymeric material has at least one particle ofthis invention.

Compositions and Methods of Use Thereof

According to another aspect of the invention there is provided a methodfor inhibition of bacteria, by contacting the bacteria with ananti-microbial particle of this invention, or a composition orpharmaceutical composition comprising the particle(s) of this invention.The term “inhibition” is referred to destruction, i.e. annihilation, ofat least 99% of the bacteria, preferably 99.9%, most preferably 99.99%of the bacteria; reduction in the growth rate of the bacteria; reductionin the size of the population of the bacteria; prevention of growth ofthe bacteria; causing irreparable damage to the bacteria; destruction ofa biofilm of such bacteria; inducing damage, short term or long term, toa part or a whole existing biofilm; preventing formation of suchbiofilm; inducing biofilm management; or bringing about any other typeof consequence which may affect such population or biofilm and imposethereto an immediate or long term damage (partial or complete).

The term “biofilm” refers to a population of biological species(bacteria) attached to a solid surface.

The terms “anti-microbial” and “anti-bacterial” are used hereininterchangeably. The quaternary ammonium and the tertiary amine groupsof this invention [—⁺N(R₁)(R₂)(R₃), —⁺NH(R¹)(R₂),—N(R¹)(R₂)—⁺N(R₁′)(R₂′)(R₃′), —⁺NH(R₁′)(R₂′) or —N(R₁′)(R₂′) (defined instructures (1) to (3))] provide the anti-microbial activity. Thequaternary ammonium's activity remains strong at any pH. Tertiary amineshave high pKa values, therefore are active at almost all pH levels (upto 10, but not higher). The tertiary amine as well as the tertiaryammonium functional groups is less likely to cause undesirable sideeffects such as irritation of soft tissue, if used in contact with skinor mucosa or if used as a pharmaceutical composition.

In a preferred embodiment, the inhibition is achieved by contacting thebacteria with a matrix containing up to 5% w/w, more preferably up to 1%particles according to this invention, or compositions comprising them.

In one embodiment, this invention further provides a composition or apharmaceutical composition comprising anti-microbial particles asreferred hereinabove. In another embodiment, thecomposition/pharmaceutical composition comprises one type of particle.In another embodiment, the composition/pharmaceutical compositioncomprises a combination of different particle types. In one embodiment,non-limiting examples for a composition/pharmaceutical composition ofthis invention are dental adhesives, bone cement, dental restorativematerials such as all types of composite based materials for fillingtooth-decay cavities, endodontic filling materials (cements and fillers)for filling the root canal space in root canal treatment, materials usedfor provisional and final tooth restorations or tooth replacement,including but not restricted to inlays, onlays, crowns, partial dentures(fixed or removable) dental implants, and permanent and temporarycements used in dentistry for various known purposes, dental andorthopedic resin based cements, sealers, composite materials, adhesivesand cements, dental restorative composites, bone cements, tooth pastes,lotions, hand-sanitizers, ointments and creams used for dermatology,wound care or in the cosmetic industry, plastic wear for medical andresearch laboratories; food packaging, mainly for dairy products andfresh meat and fish; pharmaceuticals packaging, paints for ships, thatprevent growth of biofilm or treats, breaks down and/or kills a biofilmor bacteria within, paints for bathrooms, paint for hospitals and cleanrooms; water filtration media and many others. Each possibilityrepresents a separate embodiment of this invention. In some embodiments,the particles or composition comprising thereof are used for dental andorthopedic resin based cements, sealers, composite materials, adhesinvesand cements; for dental and orthopedic metal implants and wires; forsurgical sutures; for catheters, metal surgical tools, non-surgicalmedical devices. Each possibility represents a separate embodiment ofthis invention.

In one embodiment the composition or composite of this invention is avarnish or glaze which is applied to the tooth surface, a restoration oftooth or a crown comprising the particles of this invention. In anotherembodiment the varnish or glaze provide a protective coating, lacquer;superficially polished appearance to the tooth surface, restoration orcrown of the tooth. In another embodiment, the varnish is a fluoridevarnish which is a highly concentrated form of fluoride which is appliedto the tooth's surface, as a type of topical fluoride therapy. Inanother embodiment, the aim of glazing is to seal the open pores in thesurface of a fired porcelain. Dental glazes are composed of colorlessglass powder, applied to the fired crown surface, so as to produce aglossy surface. Unglazed or trimmed porcelain may also lead toinflammation of the soft tissues it contacts.

In one embodiment, the composition/pharmaceutical composition of thisinvention is in a form selected from the group consisting of a cream, anointment, a paste, a dressing and a gel, more preferably, wherein thecomposition is formulated for topical application or administration. Inanother embodiment, the composition is intended for administration intoan oral cavity. The composition may be formulated as a tooth paste,and/or may be applied to a surface or medical device selected from thegroup consisting of: a denture cleaner, post hygienic treatment dressingor gel, mucosal adhesive paste, a dental adhesive, a dental restorativecomposite based material for filling tooth, decay cavities, a dentalrestorative endodontic filling material for filling root canal space inroot canal treatment, a dental restorative material used for provisionaland final tooth restorations or tooth replacement, a dental inlay, adental onlay, a crown, a partial denture, a complete denture, a dentalimplant and a dental implant abutment.

In one embodiment, the pharmaceutical composition further comprises atleast one pharmaceutically active ingredient. In another embodiment,non-limiting examples of pharmaceutically active ingredients includeAnalgesics, Antibiotics, Anticoagulants, Antidepressants, Anticancers,Antiepileptics, Antipsychotics, Antivirals, Sedatives and Antidiabetics.In another embodiment, non-limiting examples of Analgesics includeparacetamol, non-steroidal anti-inflammatory drugs (NSAIDs), morphineand oxycodone. In another embodiment, non-limiting examples ofAntibiotics include penicillin, cephalosporin, ciprofolxacin anderythromycin. In another embodiment, non-limiting examples ofAnticoagulants include warfarin, dabigatran, apixaban and rivaroxaban.In another embodiment, non-limiting examples of Antidepressants includesertraline, fluoxetine, citalopram and paroxetine. In anotherembodiment, non-limiting examples of Anticancers include Capecitabine,Mitomycin, Etoposide and Pembrolizumab. In another embodiment,non-limiting examples of Antiepileptics include Acetazolamide, Clobazam,Ethosuximide and lacosamide. In another embodiment, non-limitingexamples of Antipsychotics include Risperidone, Ziprasidone,Paliperidone and Lurasidone. In another embodiment, non-limitingexamples of Antivirals include amantadine, rimantadine, oseltamivir andzanamivir. In another embodiment, non-limiting examples of Sedativesinclude Alprazolam, Clorazepate, Diazepam and Estazolam. In anotherembodiment, non-limiting examples of Antidiabetics include glimepiride,gliclazide, glyburide and glipizide.

In another embodiment, the pharmaceutical composition further comprisesexcipients. In another embodiment, the excipient comprises binders,coatings, lubricants, flavors, preservatives, sweeteners, vehicles anddisintegrants. In another embodiment, non-limiting examples of bindersinclude saccharides, gelatin, polyvinylpyrolidone (PVP) and polyethyleneglycol (PEG). In another embodiment, non-limiting examples of coatingsinclude hydroxypropylmethylcellulose, polysaccharides and gelatin. Inanother embodiment, non-limiting examples of lubricants include talc,stearin, silica and magnesium stearate. In another embodiment,non-limiting examples of disintegrants include crosslinkedpolyvinylpyrolidone, crosslinked sodium carboxymethyl cellulose(croscarmellose sodium) and modified starch sodium starch glycolate.

In one embodiment, the invention is directed to a packaging compositioncomprising a thermoplastic polymer and/or hydrogel embedded withanti-microbial particles as referred hereinabove. In another embodiment,the thermoplastic polymer and/or hydrogel is embedded with a mixture oftwo or more different particles. In another embodiment, the packagingcomposition is used in the packaging of food, beverage, pharmaceuticalingredients, medical devices, surgical equipment before operation, preoperation equipment, cosmetics, and sterilized equipment/materials.

In one embodiment the packaging composition comprises a thermoplasticpolymer and/or hydrogel embedded with the particles as referredhereinabove. In another embodiment, the thermoplastic polymer ispolyvinylchloride (PVC), polyethylene, polypropylene, silicone, epoxyresin or acrylic polymers. In another embodiment, the thermoplasticpolymer is poly methylmethacrylate or polyurethane.

In another embodiment, the packaging composition further comprisesbinders, coatings, lubricants and disintegrants. In another embodiment,non-limiting examples of binders include saccharides, gelatin,polyvinylpyrolidone (PVP) and polyethylene glycol (PEG). In anotherembodiment, non-limiting examples of coatings includehydroxypropylmethylcellulose, polysaccharides and gelatin. In anotherembodiment, non-limiting examples of lubricants include talc, stearin,silica and magnesium stearate. In another embodiment, non-limitingexamples of disintegrants include crosslinked polyvinylpyrolidone,crosslinked sodium carboxymethyl cellulose (croscarmellose sodium) andmodified starch sodium starch glycolate.

In one embodiment, the packaging composition is used for packagingpharmaceutical ingredients. In another embodiment, non-limiting examplesof pharmaceutical ingredients include analgesics, antibiotics,anticoagulants, antidepressants, anti-cancers, antiepileptics,antipsychotics, antivirals, Sedatives and antidiabetics. In anotherembodiment, non-limiting examples of analgesics include paracetamol,non-steroidal anti-inflammatory drugs (NSAIDs), morphine and oxycodone.In another embodiment, non-limiting examples of antibiotics includepenicillin, cephalosporin, ciprofloxacin and erythromycin. In anotherembodiment, non-limiting examples of anticoagulants include warfarin,dabigatran, apixaban and rivaroxaban. In another embodiment,non-limiting examples of Antidepressants include sertraline, fluoxetine,citalopram and paroxetine. In another embodiment, non-limiting examplesof anti-cancers include Capecitabine, Mitomycin, Etoposide andPembrolizumab. In another embodiment, non-limiting examples ofantiepileptics include Acetazolamide, Clobazam, Ethosuximide andlacosamide. In another embodiment, non-limiting examples ofantipsychotics include Risperidone, Ziprasidone, Paliperidone andLurasidone. In another embodiment, non-limiting examples of antiviralsinclude amantadine, rimantadine, oseltamivir and zanamivir. In anotherembodiment, non-limiting examples of sedatives include Alprazolam,Clorazepate, Diazepam and Estazolam. In another embodiment, non-limitingexamples of antidiabetics include glimepiride, gliclazide, glyburide andglipizide.

In one embodiment, the packaging composition is used in the packaging offood ingredients. In another embodiment, non-limiting examples of foodingredients packaged with the packaging material of the inventioninclude fresh food, preservatives, sweeteners, color additives, flavorsand spices, nutrients, emulsifiers, binders and thickeners. In anotherembodiment, non-limiting examples of fresh food include: meat, poultry,fish, dairy products, fruits and vegetables. In another embodiment,non-limiting examples of preservatives include Ascorbic acid, citricacid, sodium benzoate, calcium propionate, sodium erythorbate, butylatedhydroxy toluene (BHT), silver, chlorhexidine, trichlozan and sodiumnitrite. In another embodiment, non-limiting examples of sweetenersinclude Sucrose (sugar), glucose, fructose, sorbitol, mannitol and cornsyrup. In another embodiment, non-limiting examples of color additivesinclude Orange B, Citrus Red No. 2, annatto extract, beta-carotene,grape skin extract, cochineal extract or carmine and paprika oleoresin.In another embodiment, non-limiting examples of flavors and spicesinclude monosodium glutamate, glycine slats, inosinic acid, isoamylacetate, and limonene and allyl hexanoate. In another embodiment,non-limiting examples of nutrients include Thiamine hydrochloride,riboflavin (Vitamin B₂), niacin, niacinamide, folate or folic acid. Inanother embodiment, non-limiting examples of emulsifiers include Soylecithin, mono- and diglycerides, egg yolks, polysorbates and sorbitanmonostearate. In another embodiment, non-limiting examples of bindersand thickeners include Gelatin, pectin, guar gum, carrageenan, xanthangum and whey.

In one embodiment, this invention provides a method for inhibiting orpreventing biofilm formation, comprising applying onto a susceptible orinfected surface or a medical device a composition of this invention.

In another embodiment, this invention provides a composition of thisinvention for use in inhibiting or preventing a biofilm formation.

In one embodiment, this invention provides a method for inhibiting orpreventing biofilm formation or growth comprising placing a medicaldevice of this invention (comprising a composition of this invention asreferred hereinabove) on the surface to be treated. In anotherembodiment, the medical device is a wound dressing.

In another embodiment, this invention provides a medical device of thisinvention for use in inhibiting or preventing biofilm formation orgrowth.

In one embodiment, this invention provides a method for inhibition ofbacteria, the method comprising the step of contacting the bacteria withthe pharmaceutical or packaging composition or composite of thisinvention.

In another embodiment, this invention provides a pharmaceutical orpackaging composition or for use in inhibiting bacteria.

In one embodiment, this invention provides a method for treating,breaking down or killing biofilm or bacteria within, comprising applyingonto a susceptible or infected surface or a medical device thepharmaceutical or packaging composition or composite of this invention.

In another embodiment, this invention provides a composite or apharmaceutical or packaging composition of this invention for use intreating, breaking down or killing biofilm or bacteria within.

Applications out of the medical field may for example be in clothing(e.g. for sports or outdoor activity; to prevent bacteria-induced sweatodor), athlete shoes or the inner part of a shoe wherein bacteria tendto collect, sportswear and clothing for outdoor activity, tooth brushesand any brush that are in contact with the human body, air and waterfilters, water treatment and distribution systems, pet cages as well asother veterinary items, etc.

In some embodiments, the anti-microbial compositions or composites ofthis invention affect annihilation of at least about 99% of thecontacted bacteria, preferably, at least about 99.99% of the contactedbacteria.

It was further surprisingly discovered that the particles withincompositions/composites/medical devices of this invention maintain highanti-microbial properties over time without leaching out and with noalteration of the properties of the hosting matrix. Such particlesdemonstrate enhanced anti-bacterial activity originating from thepresence of closely packed anti-bacterial groups on a given particle'ssurface.

Medical Devices of this Invention

In one embodiment, this invention further provides a medical devicecomprising a composition of this invention. In one embodiment,non-limiting examples for medical devices of this invention arecatheters, stents, surgical mesh, breast implants, joint replacements,artificial bones, artificial blood vessels, artificial heart valves(cardiology), artificial skin, plastic surgery implants or prostheses,intra uterine devices (gynecology), neurosurgical shunts, contact lenses(ophthalmology), intraocular lenses, ocular prosthesis, urethral stents,coating for subcutaneous (such as orthopedic or dental) implants,insulin pumps, contraceptives, pacemakers, tubing and cannulas used forintra venous infusion, tubing and cannulas used for dialysis, surgicaldrainage tubing, urinary catheters, endotracheal tubes, wound covering(dressing and adhesive bandage) and treatment (e.g. gels, ointments,pastes and creams for wound care which reduce biofilm and bacteria toaid wound healing) materials, sutures, catheters of all kinds that areinserted temporarily or permanently in blood vessels as well as theurinary system, shunt for use in brain applications, surgical gloves,tips for ear examination, statoscope ends and other elements used by themedical personnel; tooth brushes, tooth pick, dental floss, interdentaland tongue brushes, surgical sutures, metal surgical tools, non-surgicalmedical devices, dental, and orthopedic metal implants and wires andsurgical drains, syringes, trays, tips, gloves and other accessoriesused in common medical and dental procedures.

In one embodiment, this invention further provides a medical devicecomprising a dental appliance. In one embodiment, this invention furtherprovides a medical device comprising an orthodontic appliance. Thedental appliance and the orthodontal appliance comprise the particlesand composition of this invention. In some embodiments, the orthodontalappliance include an aligner for accelerating the tooth aligning, abracket, a dental attachment, a bracket auxiliary, a ligature tie, apin, a bracket slot cap, a wire, a screw, a micro-staple, cements forbracket and attachments and other orthodontic appliances, a denture, apartial denture, a dental implant, a periodontal probe, a periodontalchip, a film, or a space between teeth. In some embodiments, the dentalappliance include a mouth guard, used to prevent tooth grinding (bruxer,Bruxism), night guard, an oral device used for treatment/preventionsleep apnea, teeth guard used in sport activities.

In one embodiment, this invention further provides a trans dermalmedical device such as orthopedic external fixation screws and wiresused for bone fixations and stabilization and trans mucosal elementsused in dental implants such as healing caps, abutments (such asmultiunit), for screw retained or for cement retained dental prosthesis.

In one embodiment, this invention further provides a medical devicecomprising an endoscope (rigid and flexible), including, and not limitedto a colonoscope, gastroscope, duodenoscope, bronchoscope, cystoscope,ENT scopes, laparoscope, laryngoscope and similar instruments forexamination or treatment the inside of the patient's body, including anyparts thereof, as well as accessories and other devices used in theprocedure which either come in contact with body tissue or fluids;tubes, pumps, containers and connectors (used inside or outside thebody) through which fluids, air or gas may be pumped into or suctionedout from the patient and could become contaminated by the patient ortransfer contaminants from other patients; items such as brushes, trays,covers, tubes, connectors cabinets and bags used for reprocessing,cleaning, transporting and storing such equipment and can transmit orhost biological contaminants, as well as filters for air or water usedin dental or medical procedures, hospital surfaces (such as floors,tabletops), drapes, curtains, linen, handles and the like.

The antimicrobial property may protect the patient and the medical stafffrom cross contamination from patient to patient or from patient to theexaminer. Self-sterilizing packaging for medicines and items that enterthe operation room are also beneficial.

In one embodiment, this invention further provides processes forpreparing the medical devices comprising the composites. In anotherembodiment, the medical devices are prepared via the steps of: providinga fluid phase of the composite of this invention; shaping the fluid; andhardening of the shaped fluid, affording the desired medical device. Inanother embodiment, the medical devices are prepared via the steps of:providing a solid phase of the composite; and shaping of the solid,affording the desired medical device. In another embodiment, the shapingis accomplished via extrusion or molding. In another embodiment, fluidphase of the composite comprises melted composite or a compositedissolved in a solvent.

Another polymer material to be used in the context of this invention isresins used in dental, surgical, chirurgical and orthopedic compositematerials. In such applications, anti-microbial particles could be firstdispersed within the resin part or added simultaneously with filler orany other solid ingredients (if any). Most of these resins are acrylicor epoxy type monomers that undergo polymerization in-vivo.

The following examples are presented in order to more fully illustratethe preferred embodiments of this invention. They should in no way,however, be construed as limiting the broad scope of this invention.

EXAMPLES Example 1 Preparation of Core Particles of Amorphous SiO₂(Silica)

Silica dioxide core particles were prepared by hydrolysis of tetraalkoxysilicate under alkaline conditions. The reaction solution was preparedby mixing 9 parts by weight of ethanol, 0.4 parts of deionized water and0.1 part of ammonia, keeping the pH within the range of 10-14.Controlling the particle size and the reaction rate is achieved byadjusting the concentration of water and ammonia in the reactionsolution. 0.5 parts of tetraethyl orthosilicate (TEOS) was added to thesolution in one portion with stirring at 1,000 RPM for 1 hour. Thereaction mixture first turned opaque, followed by a white solidprecipitation, indicating the reaction endpoint and agglomeratesformation of primary particles. The particles were recovered bycentrifugation filtration, rinsing with 20 parts of deionized water anddrying using freeze drying or heating. Optionally, further surfaceactivation may be performed by shortly rinsing particles in sulfuricacid/hydrogen peroxide solution commonly known as “pirhana solution”.This last step converts most of the particles' surface into hydroxylform and promotes an efficient surface functionalization.

Example 2 Morphological Characterization of Silica Particles

Nitrogen adsorption method was used to determine the morphology ofporous silica dioxide particles by utilizing Barrett-Joyner_Halenda(BJH) model. Non-functionalized mesoporous silica dioxide particles wererinsed in Milli-Q water, dried and then degassed. Pore size was obtainedfrom the adsorption/desorption isotherm by applying BJH model. Averageparticle size measured using dynamic light scattering method. Therefore,said particles are of 186 nm in diameter and having pore size of 5.0 nm.

Example 3 Preparation of Magnetite Core Particles

Magnetite (Fe₃O₄) particles were prepared by co-precipitation of Fe²⁺and Fe³⁺ ions, from FeCl₂ (1 mol eq) and FeCl₃ (0.5 mol eq) in aqueoussolution in basic condition utilizing NH₄OH (pH˜12). Afterprecipitation, the particles recovered under constant magnetic field.Prior to functionalization, particles were rinsed in Mili-Q waterfollowed by vacuum drying. Surface activation of the obtained magnetiteparticles was performed by a short rinse of the particles in nitric acidor sulfuric acid and hydrogen peroxide solution. The last step convertedmost of particles' surface into hydroxy form allowing furtherfunctionalization of the core.

Example 4 Surface Functionalization of Inorganic Core Particles SolidSupport Method

Within the solid support method, a few stages were employed. First, thelinker 3-aminopropyltrimethoxysilane was allowed to condense ontoparticles surface (surface functionalization) via hydrolysis of methoxygroups, resulting in the attachment of the linker to the silica core(FIG. 10, step 1). Second, the attached linker was elongated, byconsecutive addition of 1,2-dichloroethane and 1,2-diaminoethane (FIG.10, steps 2 and 3). In some cases such consecutive addition was repeatedfor a few times, depending on the desired number of antimicrobialgroups. Finally, the anti-microbial active group, was grafted toresulting attached and elongated linker, via the acyl bromide moiety(FIG. 10, step 4).

Solution Method

Within the solution method, a few stages were employed. In the firststage the linker molecule was elongated by consecutive addition of1,2-dichloroethane and 1,2-diaminoethane (FIG. 11, steps 1 and 2). Insome cases such consecutive addition was repeated for a few times,depending on the desired number of antimicrobial groups. In the secondstage, the anti-microbial active group was grafted to resulting attachedand elongated linker, via the acyl bromide moiety (FIG. 11, step 3).Finally, the elongated, anti-microbial active linker was attached to thesilica core via functionalization thereof. In this stage, the linkermolecule was allowed to condense onto particles surface (surfacefunctionalization) via hydrolysis of methoxy groups, resulting in theattachment of the linker to the core (FIG. 11, step 4).

Functionalization of silica particles was performed in two stages.Initially, primary amine-functionalized silica particles were prepared.The primary amine was the functionalized by reductive amination to yielda tertiary amine comprising terpenoid groups, or alternatively aquaternary ammonium group comprising one elongated alkyl chain of 8carbons.

A pretreatment of inorganic cores (for example SiO₂, Fe₃O₄) wasessential for removing any of residual organic material such as solventor other ligands and converts the surface to active hydroxyl group thatare ready to undergo functionalization (silanization). The pretreatmentincluded rinsing the particle in 20 to 40% solution of hydrogen peroxidein sulfuric acid or alternatively in 20 to 40% of NH₄ solution insulfuric acid for at least 5 minutes at ambient conditions or atelevated temperature, preferable at least for 30 minutes at 60° C.

Polymerization of the silane groups (FIG. 8C, Mode B) versus simplesilanization (FIG. 8C, Mode A) was conducted by immersion of dryparticles in dry toluene (1 to 10 g of particles; 50 ml toluene). Excessof silane coupling agent (for example APTES) was added at ratio of atleast 10 mmol per 1 g of particles in the presence of catalytic acid(preferable acetic or hydrochloric acid). The coupling/polymerizationwas conducted at 60° C. for 1 h, then heated to 120° C. and stirredunder reflux for at least 3 h. Concentrations of silane coupling agent,acid, temperature and time during the reaction determine the mode offunctionalization (Mode A vs. Mode B) and the overall degree of surfacedensity.

Example 5 Anti-Microbial Activity of Matrix Comprising FunctionalizedSilica Particles Anti-Microbial Test Conditions—Direct Contact Test

Direct contact between bacteria and the tested materials was achieved byapplying 10 μl of bacterial suspension on each tested material sample ina set of 8 wells. The plate was incubated at a vertical position for 1 hat 37° C. During this incubation period, the suspension's liquidevaporated and a thin layer of bacteria was obtained, ensuring directcontact between the bacteria and the tested material. The plate was thenplaced horizontally and 220 μl of brain-heart infusion broth were addedto each well containing the material. All tests were done usingStapilococcus aureus (S. aureus) and Enterococcus faecalis (E. faecalis)as representative for Graham positive bacteria and Pseudomonasaeruginosa (P. aeruginosa) as representative for Graham negativebacteria.

The kinetic measurement of bacterial growth was done utilizingtemperature controlled microplate spectrophotometer (VERSAmax, MolecularDevices Corporation, Menlo Oaks Corporate Centre, Menlo Park, Calif.,USA). The microtiter plate was placed in the spectrophotometer, at 37°C. with 5 sec vortex prior to every reading. Bacterial growth wasestimated by the OD changes in each well at 650 nm every 20 minutes for24 hours.

Sample Preparation 1) Polypropylene Comprising Quaternary AmmoniumFunctionalized Silica Particles

Silica particles of an average diameter of 186 nm functionalized withquaternary dimethyl octyl ammonium were embedded in polypropylene.Samples of polymer films were prepared by hot molding of polypropyleneand the functionalized silica particles at 0, 1 and 2% wt/wt ofparticles. 5×10 mm samples of prepared films were positioned into wellsof microtitre plate touching the inside sidewalls of each well.

The anti-bacterial test results demonstrated a consistently low OD (0.1)level during the experiment for the polypropylene samples containing 1and 2% wt/wt of particles, while the polypropylene sample containing noparticles and the control sample containing S. aureus demonstrated asignificant OD increase (0.7) (FIG. 12).

Similar results were obtained in the presence of P. aeruginosa, wherethe polypropylene samples containing 2% wt/wt of particles demonstrateda low OD level (0.05) and the sample containing 1% wt/wt of particlesshowed a slightly higher OD level (0.15). In contrast, the polypropylenesample containing no particles and the control sample containing P.aeruginosa demonstrated a significant OD increase (0.7) (FIG. 13).

These results reveal the anti-microbial effect obtained by the modifiedpolypropylene substrate utilizing quaternary ammonium functionalizedsilica particles. Particles that used in this experiment had largenumber of 170 anti-microbial active functional group (170=(n₁+n₂)×m×p;n₁, n₂, m and p are defined in structure 1) grafted per nm² of thesurface of the core.

2) Poly (Methyl Methacrylate) Comprising Quaternary Amine FunctionalizedSilica Particles

Silica particles of an average diameter of 13 μm functionalized withquaternary dimethyl octyl ammonium were embedded in commerciallyavailable dental polymerizable methylmethacrylate (Unifast Trad, GCAmerica inc) at concentration of 0 and 1% wt/wt. The methylmethacrylatewas mixed in a silicone crucible at a liquid/powder ratio of 2 g/mlrespectively, in accordance to manufacturer's instructions and thenallowed to polymerize onto sidewalls of microtiter wells at 37° C. for24 hours prior to the anti-microbial test. Particles that used in thisexperiment had large number of 170 anti-microbial active functionalgroup grafted per nm² of the surface of the core (170=(n₁+n₂)×m×p; n₁,n₂, m and p are defined in structure 1).

The anti-bacterial test results demonstrated a consistently low OD (0.1)level during the experiment for the methymethacrylate (PMMA) samplescontaining 1% wt/wt of particles, while the PMMA sample containing noparticles and the control sample containing P. aeruginosa demonstrated asignificant OD increase (0.8) (FIG. 14).

Similar results were obtained in the presence of S. aureus, where PMMAsample containing 1% wt/wt of particles demonstrated a low OD level(0.1) and the sample containing no particles and the control samplecontaining S. aureus demonstrated a significant OD increase (0.8) (FIG.15).

These results reveal the anti-microbial effect obtained by the modifiedPMMA substrate utilizing quaternary ammonium functionalized silicamacro-size particles.

3) Poly (Methyl Methacrylate) Comprising Tertiary Amine FunctionalizedSilica Particles

Silica particles of an average diameter of 186 nm functionalized withdi-cinnamyl amine (tertiary amine) were embedded in commerciallyavailable dental polymerizable methylmethacrylate (Unifast Trad, GCAmerica Inc.) at concentration of 0 and 1% wt/wt. The methymethacrylatewas mixed in a silicone crucible at a liquid/powder ratio of 2 g/mlrespectively, in accordance to manufacturer's instructions and thenallowed to polymerize onto sidewalls of microtiter wells at 37° C. for24 hours prior to the anti-microbial test. Particles that were used inthis experiment had large number of 170 anti-microbial active functionalgroup grafted per nm² of the surface of the core (170=(n₁+n₂)×m×p; n₁,n₂, m and p are defined in structure 1).

The anti-bacterial test results demonstrated a consistently low OD levelduring the experiment for the methymethacrylate (PMMA) samplescontaining 1% wt/wt of particles, while the PMMA sample containing noparticles and the control sample containing P. aeruginosa demonstrated asignificant OD increase (FIG. 16).

Similar results were obtained in the presence of S. aureus, where PMMAsample containing 1% wt/wt of particles demonstrated a low OD level(0.1) and the sample containing no particles and the control samplecontaining S. aureus demonstrated a significant OD increase (0.7) (FIG.17).

These results reveal the anti-microbial effect obtained by the modifiedPMMA substrate utilizing di-terpenoid (tertiary amine) functionalizedsilica-core based particles.

4) Poly (Methyl Methacrylate) Comprising Quaternary Amine FunctionalizedMagnetite Particles

Magnetite (Fe₃O₄) particles of an average diameter of 78 nmfunctionalized with quaternary dimethyl octyl ammonium (prepared asdescribed in Example 3) were embedded in commercially available dentalpolymerizable methylmethacrylate (Unifast Trad, GC America inc) atconcentration of 0, 1 and 2% wt/wt. The PMMA was mixed in a siliconecrucible at a liquid/powder ratio of 2 g/ml respectively, in accordanceto manufacturer's instructions and then allowed to polymerize ontosidewalls of microtiter wells at 37° C. for 24 hours prior to theanti-microbial test.

The anti-bacterial test results demonstrated a consistently low OD level(0.1) during the experiment for the methymethacrylate (PMMA) samplescontaining 1 and 2% wt/wt of particles, while the PMMA sample containingno particles and the control sample containing E. faecalis demonstrateda significant OD increase (0.8) (FIG. 18).

These results reveal the anti-microbial effect obtained by the modifiedPMMA substrate utilizing quaternary ammonium functionalizedmagnetite-core based particles.

5) Poly (Methyl Methacrylate) Comprising Quaternary Amine FunctionalizedSilica Particles

Silica particles of an average diameter of 186 nm functionalized withquaternary ammonium comprising di-cinnamyl methyl substitutes (preparedas described in Example 4), were embedded in commercially availabledental polymerizable methylmethacrylate (Unifast Trad) at concentrationof 0, 2 and 3% wt/wt. The PMMA was mixed in a silicone crucible at aliquid/powder ratio of 2 g/ml respectively, in accordance tomanufacturer's instructions and then allowed to polymerize ontosidewalls of microtiter wells at 37° C. for 24 hours prior to theanti-microbial test. Both liquid and solid parts of the polymer materialwere manipulated accordingly to manufacturer's instructions and thenallowed to polymerize onto sidewalls of microtiter wells at 37° C. for24 hours prior to the anti-microbial test.

The anti-bacterial test results demonstrated a low OD (0.1) level duringthe experiment for the methymethacrylate (PMMA) samples containing 3%wt/wt of particles, and a slightly higher level for the samplecontaining 2% wt/wt of particles. In contrast, the PMMA samplecontaining no particles and the control sample containing E. faecalisdemonstrated a significant OD increase (0.7) (FIG. 19).

These results reveal the anti-microbial effect obtained by the modifiedPMMA substrate utilizing di-terpenoid quaternary ammonium functionalizedsilica-core based particles.

Example 6 Mechanical Tests of Resins Comprising Functionalized Particles

Poly methylmethacrylate (Unifast Trad) cylindrical specimens of 0.4 mmin diameter and 10 mm in length were prepared using polypropylenepipe-like molds. Specimens were allowed to polymerize at roomtemperature for 1 hour within the molds, then stored in DDW at 37° C.for 24 hours prior to testing. Each tested group contained 10 specimensof cured cement with 8% wt/wt NPs. A control group was obtained usingthe polymer specimens without functionalized particles. Compressivestrength test was carried out using universal testing machine (Instron3366, Canton, Mass.) operated at displacement speed of 1 mm/min. Datawas instantly analyzed with Merlin software which calculated thecompressive strength and the Young's modulus.

The NPs tested were marked as follows:

1) SiCial—containing 8% wt of silicadioxide particles functionalizedwith tertiary amine functional group having two cinnamyl substituentswith diameter of 186 nm (prepared as defined in Example 4).2) QPEI—containing 8% wt of dimethyl octyl quaternary ammoniumfunctionalized PEI particles of 24 nm (prepared as defined in Example5).3) A sample of unmodified poly methylmethacrylate (PMMA) resin was usedas a control.

The results demonstrated relatively high stability of the modifiedacrylate resin comprising the silica based particles under stressconditions. The compressive strength of unmodified PMMA, SiCial and QPEIare 56.61, 78.79 and 0.43 MPa respectively. The embedment of silicafunctionalized antibacterial particles did not jeopardize the mechanicalproperties of the resin, and appeared to be advantageous in terms ofstress-stability in comparison to the polymeric functionalized resin(QPEI) (FIG. 20B).

Example 7 Antibacterial Test of Resins Comprising FunctionalizedParticles

The samples described on Example 7 were tested for their antibacterialactivity by direct contact test as described herein above (Example 6).

The results demonstrate the potent antibacterial effect of the modifiedresins due to the embedment of the functionalized silica-based andPEI-based particles compared with the unmodified resin control sampleand the natural growth of bacteria as depicted in the presence of E.faecalis (FIG. 21A) and S. aureus (FIG. 21B).

Example 8 Antibacterial Test by Imprint Method

Three glass slides were coated utilizing spraying of a solutioncontaining functionalized silica based particles onto the hydroxylatedglass surface. The silane group anchored the functionalized particles tothe slide upon hydrolysis of the leaving groups and the slides werefurther dried at elevated temperature to allow complete condensation ofthe particles onto to the surface. The glasses were marked asfollows: 1) dimethylamine functionalized silica particles; 2) tertiaryamine with two cinnamyl groups functionalized silica particles.

S. aureus suspension was applied onto each functionalized slide in ahomogeneous manner. The slides were placed in contact with blood agarpetri dish facing towards the agar for 15 minutes. Subsequently, theslides were removed and the petri dishes were kept in 37° C. for 24 toallow formation of colonies.

The results revealed that no colonies were formed onto the petri dishwhich came in contact with functionalized slide 2, demonstrating theadvantageous antibacterial activity of the tertiary amine comprising twocinnamyl groups (FIG. 22).

Example 9 Determination of the Loading Degree of Anti-Bacterial ActiveGroups onto the Core

FIG. 23 presents a scheme of the different methods to determine the loadconcentration of the anti-microbial group onto the core.

Method 1—degree of amine loading onto particle's surface. 1.0 g of dryamine-functionalized silica particles powder having 180 nm diameter wasimmersed in 20 ml of dry toluene. Then 0.1 g (1.9 mmol) ofFluorenylmethyloxycarbonyl (Fmoc) chloride were added. The mixture wasreacted at 60° C. under continuous stirring for 12 hours. Resultingparticles were filtered and rinsed 3 times with 5 ml ofN-Methyl-2-pyrrolidone (NMP), then 3 times with 5 ml of diethyl etherand then dried in-vacuo. Detachment of Fmoc was performed by immersing0.01 g of Fmoc-labeled particles in 2 ml of 20% by volume solution ofpiperidine in NMP and shaked for 30 min followed by filtration ofsolvent. This procedure repeated once more and both solutions werecombined (to a total of 4 ml solution). Concentration of Fmoc insolution was determined using light absorbance in spectrophotometer at301 nm and calculated in accordance to Beer's law A=EbC, where A isabsorbance, E is molar absorption constant (6300 cm⁻¹M⁻¹), b is pathwaylength (lcm) and C is molar concentration. Prior to spectrometryreadings, solution was diluted at 1:100 ratio in NMP.

Results: A=1.1, therefore C=100×(1.7×10⁻⁴)M=0.017M. Therefore,N(moles)=0.017M×0.004 L=6.98×10⁻⁵ moles. Total loading is therefore6.98×10⁻⁵ mol/0.01 g=0.007 moles/gr. Assuming perfect sphere geometry ofparticles, the shell surface area of single particles is 102000 nm² andparticle average volume is 3050000 nm³. Particles density calculatedusing Archimedes method is 2.5 g/(1×10²¹ nm³), giving a singleparticle's mass of 7.6×10⁻¹⁶ g. Therefore, the loading of functionalgroups is ((7.6×10⁻¹⁶ g)×(0.007 moles/g))/102000 nm²=5.2×10⁻²³moles/nm², which is approximately 31 amine/ammonium per nm².

Method 2—degree of functional tertiary amines substituted with twocinnamyl groups. 0.001 g of 186 nm silica particles functionalized withdi-cinnamyl amines were immersed in 100 ml of absolute ethanol.Spectrophotometric reading were taken at the wavelength of 327 nm.E(cinnamaldehyde)=25118 cm⁻¹M⁻¹. All calculations were performed asdescribed in Method 1.

Results: A=1.5, therefore total tertiary amines count is 6×10⁻⁶ moles,which is 3.0×10⁻³ moles/g.

Therefore the functional groups loading is approximately 13amine/ammonium per nm².

Both methods are applicable for all kinds of inorganic and organic coreparticles, whereas for organic particles (polymeric particles) the Fmocfunctionalization is performed after the cross-linking step.

TABLE 1 Antibacterial activity dependency of polmethylmethacrylatemodified particles of the invention as a function of functional groupsdensity loaded onto particle surface. All experiments were performed asin examples 4 and 6. Surface Inhibition of P. Inhibition of densityaerginosa S. aureus Particle (units/nm²) (in Logs) (in Logs) SiO₂ core13 4 5 Quaternary ammonium 31 6 6 (octyl dimethyl 174 6 6 ammonium)func. SiO₂ core 13 3 3 di-cinnamylamine func. 31 3 5 174 5 6 Fe₃O₄ core13 2 3 Quaternary ammonium 60 3 4 (octyl dimethyl 130 5 5 ammonium)func. Fe₃O₄ core 13 0 2 di-cinnamylamine func. 60 3 4 130 4 5 PEI core12 4 5 Quaternary ammonium 120 5 6 (octyl dimethyl 230 5 6 ammonium)func. PEI core 12 3 4 di-cinnamylamine func. 120 5 5 230 5 6

As shown in the above table, the polmethylmethacrylate modifiedparticles of the invention showed antibacterial activity for bothinorganic and organic cores. The denser functional groups are packedonto particle surface, the stronger antibacterial activity against bothtested organisms, for both organic and inorganic cores and for bothquaternary ammonium salts and tertiary amines (terpenoids). Such denserpacking is found as the number of anti-microbial active groups per oneanti-microbial active part increases; for example, first (top) entry ineach inorganic core has a ratio of only one anti-microbial active groupper one anti-microbial active part, whereas other entries for theinorganic cores comprise higher ratio and those first entries have thelowest exhibited anti-bacterial activity.

Example 10 Activity of Silica Based Particles of this Invention

Four types of SiO₂ based particles were added to soft paraffin atconcentration of 2% wt and dispersed until homogeneous paste was formed,while using ceramic pestle and crucible. Samples prepared according toexample 4 and were marked as 2% Silicadioxide-di-cinnamylamine forparticles having tertiary amine functional groups with two cinnamylsubstituents, 2% Silicadioxide-quaternary ammonium for particles havingone octyl and two methyls attached to quaternary nitrogen, 2% QPEI forquaternary ammonium polyethyleneimine, 2% Silicadioxide dimethylaminofor samples having tertiary amine of two methylenes on the nitrogen and“E. faecalis” for control of paraffin-only group. Direct contact test(DCT) was performed for treated gauze pads with each one of paraffinsamples. The results (FIG. 24) demonstrate strong inhibition of bacteriagrowth for all test samples excluding the dimethylamino variation.Specifically, the activity of terpenoids substituent onto tertiary aminefunctionality is surprising, due to their immobilization unlike knownantimicrobial activity of free terpenoids.

Example 11 An Antibacterial Toothpaste Comprising Silica Based Particlesof this

Composition of antibacterial toothpaste: glycerol, water, sorbitol,sodium lauryl sarcosine, hydrated silica, titanium dioxide andantibacterial particles. The antibacterial particles comprise SiO₂particles which is commonly used in commercial toothpaste, where some ofthe particles are modified by covalently binding antibacterial groups.The antibacterial groups may be quaternary ammonium and tertiary aminehaving two cinnamyl groups or having tertiary amines with two citralgroups. Below are shown results of a toothpaste formulation containing5% wt of antibacterial SiO₂ particles having tertiary amine with twocinnamyl groups.

Surface retention experiment: Herein are presented results of particlesretention onto glass surfaces examined by simulation of tooth brushingprocedure during 1 minute with three compositions of toothpaste: A:commercially available toothpaste (control); B: the toothpastecomposition as presented above, without antibacterial particles(control) and C: proposed toothpaste with antibacterial particlesretention onto glass slides. After brushing, slides rinsed with sameamount of water in same manner. Retention examined visually (FIG. 25).The commercial toothpaste (Colgate® total) and the toothpasteformulation (with the composition as described above) withnon-functionalized SiO₂ particles show no visible retention to glasssurface. The toothpaste formulation with 5% wt of antibacterialparticles (SiO₂ with tertiary amine having two cinnamyl groups) of thecurrent invention exhibits significant and visible retention to glasssurface.

Antibacterial activity experiment: antibacterial activity of proposedtoothpaste was examined by dispersing 10 μl of S. mutars (˜10⁶ viablecells) within total volume 220 μl of phosphate buffer saline (PBS) andproposed toothpaste. In this experiment, toothpaste formulation withantibacterial particles was tested, at the following finalconcentrations (% wt.): 0, 0.25, 0.5, 1 and 2. Each sample performed in8 repetitions in 96 well plate. Bacteria growth monitored by readingoptical density at 650 nm while incubating at 37° C. (FIG. 26). Theantibacterial activity is proportional to particles concentration(dose-dependent effect). At concentration of 2% wt. there wasn't anysingle bacteria cell which survived out of the 10⁶ incubated viablebacteria cells.

Example 12 Contact Lenses Comprising Silica Based Particles of thisInvention

A contact lenses composition comprising antibacterial SiO₂ particleswith tertiary amine having two cinnamyl groups which are incorporatedinto polymethylmethacrylate at final concentration of 2% wt wereprepared. The polymerization of the polymethylmethacrylate was done inthe following method: 48 g of methyl methacrylate monomer were mixedwith 1 g of benzoyl peroxide in glass beaker using overhead stirrer at500 rpm. until complete dissolution of peroxide. In parallel, 50 g ofmethylmethacrylate were mixed with 1 g of dihydroxyethyl p-toluidineuntil complete dissolution. Into the methylmethacrylate/dihyhdroxyethyland p-toluidine solution, 2 g of SiO₂ particles having tertiary aminewith two cinnamyl groups were added and dispersed using high-shearhomogenizer at 3000 rpm until homogeneous solution was obtained. Thenboth solutions were mixed and allowed to be polymerized onto sidewallsof 96 well plate.

Antibacterial activity experiment: direct contact test (DCT) wasperformed using E. faecalis as test bacteria at 37° C. during 24 hours.FIG. 22 shows that in the present experiment the tertiary amine was moreantibacterially active than quaternary ammonium when imbedded intopolymethylmethacrylate in the same concentrations.

Example 13 Bone Cement Comprising Silica Based Particles of thisInvention

Bone cement is used in orthopedics for fixation of implants duringsurgery operations. Bone cement composition: this cement composition isbased on liquid monomer methylmethacrylate solution with initiators andsolid pre-polymerized polymethylmethacrylate with initiators asactivators, as shown above for the contact lenses.

Antibacterial activity experiment: the silica based antibacterialparticles of the current invention were added to a solid part ofcommercially available bone cement. Three samples have been tested forantibacterial activity: (I): SiO₂ particles having tertiary amine withtwo cinnamyl groups, (II): SiO₂ particles with quaternary ammonium,wherein the overall concentration of particles in each sample aftermixing with liquid part of bone cement was 2% wt and (III) unmodifiedbone cement as control in this experiment. Samples of bond cement,unmodified and modified with antibacterial particles—were applied ontosidewalls of 96 wells plate and DCT protocol was performed with S.aureus as test bacteria. FIG. 22 shows that out of 10⁶ bacteria cells,there wasn't any single bacteria cell that grew on the surface of bonecement containing 2% wt. of silica based antibacterial particles of thecurrent invention.

Example 14 Antibacterial Activity of the Silica Based AntibacterialParticles of the Current Invention in a Water Filtration Media

1 g of chloromethyl-polystyrene beads (Merrifield resin) was dispersedwithin 50 ml of dichloromethane. 1 g of SiO₂ particles having tertiaryamine with two citral groups was dispersed in 10 ml of dichloromethaneusing high shear homogenizer at 3000 rpm until homogeneous suspensionwas obtained. Both solutions were combined and stirred for 72 h at roomtemperature. Subsequently, modified beads with antibacterial particleswere rinsed 5 times with 20 ml of DCM, then twice with 20 ml of diethylether and eventually were dried under vacuum overnight.

Antibacterial activity experiment: antibacterial test was performed inbrain heart infusion (BHI) suspension of the modified beads to study theeffect on S. aureus bacteria. 220 μl of BHI suspension with variableconcentration of modified beads were poured into wells of 96 wellsplate, with 8 wells for each concentration. Subsequently, 10 ul ofbacteria (10⁶ viable cells) were added into each tested well and lightabsorbance was measured at 650 nm each 20 minutes for 24 h. During theexperiment, each plate with the sample was kept at 37° C. and shaked for5 sec before each reading. As shown in FIG. 24, partial antibacterialactivity is obtained for samples with 1% wt, followed by stronger effectfor samples with 2% wt and complete bacteria inhibition at 5% wt.

Example 15 Antibacterial Activity of Silica Based AntibacterialParticles of the Current Invention with Tertiary Amine with 2 CinnamylGroups or Quaternary Ammonium Various Surface Concentration ofFunctional Groups Per Square Nanometer

TABLE 2 antibacterial activity of polymethylmethacrylate modified withSiO₂ particles having tertiary amine with two cinnamyl groups or withSiO₂ particles having quaternary ammonium groups. Number of functionalS. mutans E. faecalis groups per reduction in reduction in square DirectContact Direct Contact Entry nanometer Test (log₁₀) Test (log₁₀) 1 SiO₂with quaternary 0.1-0.4 3 4 ammonium 2 SiO₂ with quaternary  6-10 >6 >6ammonium 3 SiO₂ with tertiary amine 0.1-0.4 2 4 with 2 cinnamyl groups 4SiO₂ with tertiary amine  6-10 >4 >6 with 2 cinnamyl groups

Table 2 demonstrates the relation between the number of functionalgroups onto silica particle and the antibacterial activity against twoselected bacteria. Entries 1 and 3 has a ratio of only oneanti-microbial active group per one anti-microbial active part, whereasother comprise higher ratios. In addition, shown the differences betweenquaternary ammonium functionality and tertiary amines with two cinnamylgroups. It is concluded that (i) the number of functional groups isproportional to the ability of the particles to inhibit bacteria growthand (ii) quaternary ammonium functionality demonstrate strongest potencyto inhibit bacteria growth than tertiary amines with 2 cinnamyl groups.

Example 16 Dental Restorative Composite of this Invention

Typical dental restorative composite was prepared by mixing thefollowing components (weight % in brackets):

Bis-GMA (bisphenol A-glycidyl methacrylate) (10% wt.);UDMA (urethane dimethacrylate) (5% wt.);TEGDMA (triethyleneglycol dimethacrylate) (5% wt.);

Camphorquinine (1% wt.);

Ethyl-4-dimethylamino benzoate (EDMAB) (1% wt.);Fumed silica (5% wt.);Silanated glass filler (73% wt.); andanti-microbial particles (2% wt of the above composition)

Example 17 Inhibition of E. faecalis Bacteria Using Composites of thisInvention

A composite of anti-microbial quaternary polyethylene imine (QPEI)particles in silicone polymer was prepared according to the following:two-part room temperature vulcanization silicone material was used asmodel silicone material used in manufacturing of silicone medicaldevices, such as breast implants and Foley catheters. Unmodifiedsilicone polymer was used as reference (marked as “silicone”). Thesilicone precursor was polymerized by pressing with a flat plastic sheetagainst flat plastic surface which QPEI mixed with the two componentswere applied to. Obtained silicone sheets were cut to 5×15 mm specimensand placed onto sidewalls of 96-weels plate. Direct contact test (DCT)performed against E. faecalis. As shown in Error! Reference source notfound., full inhibition of bacteria grow obtained at 1% wt/wt of QPEIparticles.

Example 18 Comparing Antibacterial Activity of Composites ComprisingParticles with Different Number of Monomeric Units in the Anti-MicrobialActive Part

Anti-microbial particles [silica core functionalized with a methyl octylammonium quaternary ammonium groups, wherein n=1-3 (i.e the number ofmonomeric units per anti-microbial active unit is between 1 to 3)wherein the number of anti-microbial active groups is 174 (structure 1;(n₁+n₂)×m×p=174) per one sq. nm (nm²) of the core surface] were embeddedin commercially available dental polymerizable methylmethacrylate(Unifast Trad, GC America inc) at concentration of 0-2% wt/wt. Themethylmethacrylate was mixed in a silicone crucible at a liquid/powderratio of 2 g/ml respectively, in accordance to manufacturer'sinstructions and then allowed to polymerize onto sidewalls of microtiterwells at 37° C. for 24 hours prior to the anti-microbial test.

The anti-bacterial test (direct contact test, see example 5) results(FIG. 24) demonstrated that increasing “n” leads to higheranti-bacterial activity (reduced OD of E. faecalis) and the most potentantibacterial effect was achieved when n=3.

Example 19 Antibacterial Activity of Composites Comprising Silica-CoreBased Particles with Tertiary Amine Bearing Two Cinnamaldehyde Groups(SNP-Cial)

Silica-core based particles functionalized with tertiary amine bearingtwo cinnamaldehyde groups (SNP-Cial, FIG. 25) were embedded incommercially available dental polymerizable methylmethacrylate (UnifastTrad, GC America inc). The methylmethacrylate was mixed in a siliconecrucible at a liquid/powder ratio of 2 g/ml respectively, in accordanceto manufacturer's instructions and then allowed to polymerize ontosidewalls of microtiter wells at 37° C. for 24 hours prior to theanti-microbial test. The anti-bacterial test (direct contact test, seeexample 5) results (FIG. 25) demonstrated the anti-bacterial activity(reduced OD of E. faecalis) of the composition compared to the controlin the absence of the anti-microbial particles.

Example 20 Poloxamer Hydrogel Composites Comprising Silica Nanoparticlesof this Invention

The hydrogel was prepared by reacting poly(ethylene glycol) diglycidylether with diethylenetriamine. Immediately after mixing of bothreactants, 2QSi particles [=silica core functionalized with a methyloctyl ammonium quaternary ammonium groups, wherein the number ofmonomeric units per anti-microbial active unit is 2 [m=2; Structure 1)]were introduced and mixed until uniform suspension obtained. This blendwas poured onto flat mold and left to dry at 37° C. for 48 hours tocomplete polymerization. Subsequently, the thin film of the polymer wasdipped in deionized water allowing it to absorb moisture.

The DCT protocol (example 5) was used to evaluate the antibacterialactivity of modified hydrogel with 2QSi, as presented in table 3.

TABLE 3 anti-bacterial activity of poloxamer hydrogel compositescomprising 2QSi particles against E. faecalis Composite Bacteriainhibition (Log₁₀) Poloxamer-based hydrogel + 1.5% 2QSi >7Poloxamer-based hydrogel + 1.0% 2QSi 3 Poloxamer-based hydrogel + 0.5%2QSi 0.5 Poloxamer-based hydrogel (control) 0As shown in the table, anti-bacterial activity (against E. faecalis)increased as the 2QSi particles concentration within the poloxamercomposite was increased.

Example 21 Alginate Hydrogel Composites Comprising Silica-Core BasedParticles of this Invention

2QSi particles [=silica core functionalized with a methyl octyl ammoniumquaternary ammonium groups, wherein the number of monomeric units peranti-microbial active unit is 2 [m=2; Structure 1)] were incorporatedinto alginate hydrogel by premixing dry alginate powder with 2QSiparticles. Subsequently, sufficient amount of water was added and thecompound was mixed until homogeneous paste was formed.

The hydrogel was allowed to dry onto sidewalls of DCT plates andantibacterial activity was evaluated in accordance to the DCT protocol(example 5), as presented in table 4.

TABLE 4 anti-bacterial activity of alginate hydrogel compositescomprising 2QSi particles against E. faecalis Composite Bacteriainhibition (Log₁₀) Alginate-based hydrogel + 2.0% 2QSi >7 Alginate-basedhydrogel + 1.0% 2QSi 5 Alginate-based hydrogel (control) 0As shown in the table, anti-bacterial activity (against E. faecalis)increased as the 2QSi particles concentration within the alginatecomposite was increased.

Example 22 Activity in Sub-Cutaneous Implants In-Vivo

Design: The antibacterial activity of 2QSi-POSS particles [=POSS corefunctionalized with a methyl octyl ammonium quaternary ammonium groups,wherein the number of monomeric units per anti-microbial active unit is2 [m=2; Structure 1)] incorporated in silicone implants at 2% w/w,implanted subcutaneous was tested. POSS particles having quaternaryammonium functionality with n=2 were incorporated into silicone rodsthat were implanted in the back of mice on one (right) side of thespine, and identical rods without particles were implanted on theopposite (left) side of the spine as controls. The implants wereinoculated with 10 μl of 10⁸/ml E. faecalis either one ex-vivo (beforeimplantation) (Group A, n=10) or 8 times in 2-day intervals in-situ(starting 1 week after implantation, to allow for recovery, Group B,n=4). After explanation, the implants were vortexed and rinsed to removefree (planktonic) bacteria and then rolled on Agar plate to assessbiofilm presence on the implant by CFU count (stamp test).

Results: In group A (inoculated ex-vivo), among 9/10 animals availablefor explantation and analysis, none of the particle-containing implantshad biofilm on the stamp test (zero CFU), compared to 6 control(no-particles) implants who had significant growth, 2 with minor growthand 1 with no growth. Similarly, no loosely bound bacteria were detectedin the vortexed suspension from the test implants, vs. 1.5×10³ recoveredfrom the control implants. In group B (inoculated in-situ), stamp testshowed no biofilm in 2 animals on both test and control implants, whilein the 2 other animals there was extensive growth on the controlimplants vs. no growth on the test implant. Results are summarized inTable 5 below.

These results indicate that the antibacterial particles can preventbiofilm growth and significantly reduce overall number of bacteria onsilicone subcutaneous implants.

TABLE 5 Test Implant Control Implant Groups (with particles) (noparticles) A Ex-vivo inoculation Extensive growth 0 6 Minor growth 0 2No growth 9 1 B In-situ inoculation Extensive growth 0 2 Minor growth 11 No growth 3 1

While certain features of this invention have been illustrated anddescribed herein, many modifications, substitutions, changes, andequivalents will now occur to those of ordinary skill in the art. It is,therefore, to be understood that the appended claims are intended tocover all such modifications and changes as fall within the true spiritof this invention.

What is claimed is:
 1. A dental restorative composite comprising (i) apolymeric material and (ii) an anti-microbial particles which arerepresented by structure (1):

wherein the core is an inorganic material; L1 is a first linker or abond; L2 is a second linker; L3 is a third linker or a bond; R1 and R1′are each independently unsubstituted C1-C10 alkyl, terpenoid,cycloalkyl, aryl, heterocycle, alkenyl, alkynyl or any combinationthereof, R2 and R2′ are each independently unsubstituted C1-C10 alkyl,terpenoid, cycloalkyl, aryl, heterocycle, alkenyl, alkynyl or anycombination thereof, R3 and R3′ are each independently not present,hydrogen, C4-C8 alkyl, terpenoid moiety, cycloalkyl, aryl, heterocycle,alkenyl, or alkynyl; wherein if R3 or R3′ are not present, the nitrogenis not charged; X1 and X2 is each independently a bond, alkylene,alkenylene, or alkynylene; p defines the density of anti-microbialactive unit per one square nm (nm²) of the core surface, wherein saiddensity is of between 1-100 anti-microbial active units per one squarenm (nm²) of the core surface of the particle; n₁ an integer 0 or 1; n₂is an integer 0 or 1; m is an integer 1; and wherein said composite isused in dental applications.
 2. The composite of claim 1, wherein theparticle is represented by Structure (4):

wherein X′ is nothing or hydrogen; wherein at least one of R₁, R₂, R₃ ishydrophobic having at least four carbons.
 3. The composite of claim 1,wherein the particles are dispersed in the polymeric material.
 4. Thecomposite of claim 1, wherein the core of the particles comprisessilica.
 5. The composite of claim 4, wherein the silica is selected fromthe group consisting of amorphous silica, dense silica, aerogel silica,porous silica, mesoporous silica and fumed silica.
 6. The composite ofclaim 1, wherein the inorganic polymer is selected from the groupconsisting of silicone polymers ceramics, metals, and combinationsthereof.
 7. The composite of claim 1, wherein the inorganic core ispolyhedral oligomeric silsesquioxane (POSS).
 8. The composite of claim1, wherein the weight ratio of the anti-microbial particles which arerepresented by structure (1) to the polymeric material is between0.25-5%.
 9. The composite of claim 1, wherein the particles are amixture of different particles.
 10. The composite of claim 1, whereinsaid composition is capable of filling of tooth decay cavities, is adental restorative endodontic filling material for filling root canalspace in root canal treatment, or is selected from the group consistingof a dental restorative material intended for provisional and finaltooth restorations or tooth replacement, a dental inlay, a dental onlay,a crown, a partial denture, a complete denture, a dental implant, adental implant abutment, and a cement intended for permanently cementingcrowns bridges, onlays, partial dentures and orthodontic appliances ontotooth enamel and dentin.
 11. The composite of claim 1, wherein theanti-microbial active groups have a surface density of 1-20anti-microbial groups per 1 square nm of the core surface.
 12. Thecomposite claim 1, wherein the anti-microbial active groups have asurface density of 50-100 anti-microbial groups per square nm of thecore surface.
 13. The composite of claim 1, wherein said composite isprepared by mixing Bis-GMA (bisphenol A-glycidyl methacrylate); UDMA(urethane dimethacrylate); TEGDMA (triethyleneglycol dimethacrylate);Camphorquinine; Ethyl-4-dimethylamino benzoate (EDMAB); Fumed silica;Silanated glass fille; and the anti-microbial particles represented bystructure (1).
 14. The composite of claim 13, wherein the components ofsaid mixture are in the following amounts: Bis-GMA: 10% wt.; UDMA: 5%wt.; TEGDMA: 5% wt.; Camphorquinine: 1% wt.; EDMAB: 1% wt.; Fumedsilica: 5% wt.; Silanated glass filler: 73% wt.; and the anti-microbialparticles represented by structure (1): 2% wt.